Soybean plant and seed corresponding to transgenic event mon87712 and methods for detection thereof

ABSTRACT

The present invention provides a transgenic soybean comprising event MON87712 that exhibits increased yield. The invention also provides cells, plant parts, seeds, plants, commodity products related to the event, and DNA molecules that are unique to the event and were created by the insertion of transgenic DNA into the genome of a soybean plant. The invention further provides methods for detecting the presence of said soybean event nucleotide sequences in a sample, probes and primers for use in detecting nucleotide sequences that are diagnostic for the presence of said soybean event.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application Nos.61/392,267 and 61/393,448, filed on Oct. 12, 2010 and Oct. 15, 2010,herein incorporated by reference in their entirety.

INCORPORATION OF SEQUENCE LISTING

The sequence listing that is contained in the file named“MONS299WO.txt”, which is 26.3 kilobytes as measured in MicrosoftWindows operating system and was created on 5 Oct. 2011, is filedelectronically herewith and incorporated herein by reference.

FIELD OF THE INVENTION

The present invention relates to transgenic soybean event MON87712 andplants comprising the event that exhibit increased yield. The inventionalso provides cells, plant parts, seeds, plants, commodity productsrelated to the event, and DNA molecules that are unique to the event andwere created by the insertion of transgenic DNA into the genome of asoybean plant. The invention further provides methods for detecting thepresence of said soybean event nucleotide sequences in a sample, probesand primers for use in detecting nucleotide sequences that arediagnostic for the presence of said soybean event.

BACKGROUND OF THE INVENTION

Soybean (Glycine max) is an important crop and is a primary food sourcein many areas of the world. The methods of biotechnology have beenapplied to soybean for improvement of agronomic traits and the qualityof the product. One such agronomic trait is increased yield.

Increased yield may be achieved in transgenic plants by the expressionof a transgene capable of providing such increased yield. The expressionof foreign genes in plants is known to be influenced by many factors,such as the regulatory elements used in the transgene cassette, thechromosomal location of the transgene insert, the proximity of anyendogenous regulatory elements close to the transgene insertion site,and environmental factors such as light and temperature. For example, ithas been observed that there may be a wide variation in the overalllevel of transgene expression or in the spatial or temporal pattern oftransgene expression between similarly-produced events. For this reason,it is often necessary to screen hundreds of independent transformationevents in order to ultimately identify one event useful for commercialagricultural purposes. Such an event, once identified as having thedesired transgene expression, molecular characteristics and the improvedtrait, may then be used for introgressing the improved trait into othergenetic backgrounds using plant breeding methods. The resulting progenywould contain the transgenic event and would therefore have thetransgene expression characteristics for that trait of the originaltransformant. This may be used to produce a number of different cropvarieties that comprise the improved trait and are suitably adapted tospecific local growing conditions.

SUMMARY OF THE INVENTION

The present invention provides transgenic soybean plants and seedscomprising event MON87712, a representative seed sample of which havebeen deposited with American Type Culture Collection (ATCC) under theAccession No. PTA-10296. Plants comprising the event exhibit increasedyield.

The invention provides a plant, seed, cell, progeny plant, or plant partcomprising the event and commodity products derived from a plant, cell,plant part, or seed comprising event MON87712. The invention thusprovides a plant, seed, cell, progeny plant, plant part, or commodityproduct comprising a DNA molecule having a nucleotide sequence selectedfrom the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3,SEQ ID NO: 4, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 6, andcomplements thereof. The invention provides a plant, seed, cell, progenyplant, or plant part comprising a recombinant DNA molecule that producesan amplicon comprising a DNA molecule of the invention, for instance ina DNA amplification method. The plant parts of the soybean comprisingevent MON87712 of the present invention include, but are not limited topollen, ovule, flower, shoot, root, stem, leaf, pod and seed. Novelgenetic compositions contained in the genome of a plant comprising eventMON87712 and products made from a plant comprising event MON87712 suchas whole or processed seed, animal feed, oil, meal, flour, foodproducts, protein supplements, fuel products and biomass from whichsoybean plants comprising MON87712 have been harvested are aspects ofthis invention.

The invention provides DNA molecules related to event MON87712. TheseDNA molecules may comprise nucleotide sequences representing or derivedfrom the junction of the transgene insertion and flanking genomic DNA ofevent MON87712, and/or a region of the genomic DNA flanking the insertedDNA, and/or a region of the integrated transgenic DNA flanking theinsertion site, and/or a region of the integrated transgenic expressioncassette, and/or a contiguous sequence of any of these regions.

In one embodiment, the invention provides a DNA molecule comprising anucleotide sequence selected from the group consisting of SEQ ID NO: 1,SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 7, SEQ ID NO: 8 andSEQ ID NO: 6, and complements thereof. In another embodiment, the DNAmolecule may comprise a polynucleotide having a sequence comprising fromat least about 51-100 consecutive nucleotides of a sequence selectedfrom the group consisting of SEQ ID NO: 7 and SEQ ID NO: 8 or apolynucleotide molecule having a sequence with at least 90% identity toSEQ ID NO: 6. In yet another embodiment, the invention provides a DNAmolecule comprising SEQ ID NO: 1 and SEQ ID NO: 2. According to stillanother embodiment, there are no more than 5000 consecutive nucleotidesbetween the 3′ end of SEQ ID NO: 1 and the 5′ end of SEQ ID NO: 2, forinstance, no more than 4500, 4000, 3500, 3000, 2500, 2250, 2100, 2050,2000, or 1992 consecutive nucleotides between the 3′ end of SEQ ID NO: 1and the 5′ end of SEQ ID NO: 2. According to yet another embodiment,there are no more than 5000 consecutive nucleotides between the 3′ endof SEQ ID NO: 7 and the 5′ end of SEQ ID NO: 8, for instance, no morethan 4500, 4000, 3500, 3000, 2500, 2250, 2100, 2050, 2000, or 1914consecutive nucleotides between the 3′ end of SEQ ID NO: 7 and the 5′end of SEQ ID NO: 8. In a further embodiment, the invention provides aDNA molecule comprising SEQ ID NO: 1 or SEQ ID NO: 2 and furthercomprising a BBX32 coding sequence. In some embodiments, the BBX32coding sequence is operably or genetically linked to SEQ ID NO: 1 or SEQID NO: 2. In other embodiments, the BBX32 coding sequence is flanked bySEQ ID NO: 1 and SEQ ID NO: 2.

The invention also provides DNA molecules useful as primers and probesdiagnostic for the event. Plants, cells, plant parts, commodityproducts, progeny, and seeds comprising these molecules are provided.

According to one aspect of the invention, compositions and methods areprovided for detecting the presence of the transgene/genomic insertionregion from a novel soybean plant comprising MON87712. DNA sequences areprovided that comprise at least one junction sequence of MON87712selected from the group consisting of SEQ ID NO: 1 (corresponding topositions 3495 through 3516 of SEQ ID NO: 6, FIG. 1 [F]), SEQ ID NO: 2(corresponding to positions 5509 through 5530 of SEQ ID NO: 6, FIG. 1[F]), SEQ ID NO: 7 (corresponding to positions 3456 through 3555 of SEQID NO: 6), SEQ ID NO: 8 (corresponding to positions 5470 through 5569 ofSEQ ID NO: 6), and complements thereof; wherein a junction sequence is anucleotide sequence that spans the point at which heterologous DNAinserted into the genome is linked to the soybean cell genomic DNA anddetection of this sequence in a biological sample containing soybean DNAis diagnostic for the presence of the soybean event MON87712 DNA in saidsample (FIG. 1). A soybean event MON87712 and soybean seed comprisingthese DNA molecules is an aspect of this invention.

According to another aspect of the invention, two DNA molecules areprovided for use in a DNA detection method, wherein the DNA moleculesfunction as DNA primers when used together in an amplification reactionwith a template derived from event MON87712 to produce an amplicondiagnostic for event MON87712 DNA in a sample. In one embodiment, afirst and second DNA molecules comprise a polynucleotide molecule havinga nucleotide sequence of sufficient length of consecutive nucleotides ofSEQ ID NO: 6, or a complement thereof.

In another embodiment, the first DNA molecule comprises a polynucleotidehaving a nucleotide sequence of sufficient length of consecutivepolynucleotide of any portion of the transgene region of the DNAmolecule of SEQ ID NO: 3 or SEQ ID NO: 5 and the second DNA moleculecomprises a polynucleotide of similar length of any portion of a 5′flanking soybean genomic DNA region of SEQ ID NO: 3. Any ampliconproduced by DNA primers homologous or complementary to any portion ofSEQ ID NO: 3 and SEQ ID NO: 5, and any amplicon that comprises SEQ IDNO: 1 or at least 51 consecutive nucleotides of SEQ ID NO: 7 is anaspect of the invention.

According to another aspect of the invention, the first DNA moleculecomprises a polynucleotide having a nucleotide sequence of sufficientlength of consecutive polynucleotide of any portion of the transgeneregion of the DNA molecule of SEQ ID NO: 4 or SEQ ID NO: 5 and thesecond DNA molecule comprises a polynucleotide of similar length of anyportion of a 3′ flanking soybean genomic DNA of SEQ ID NO: 4. Anyamplicons produced by DNA primers homologous or complementary to anyportion of SEQ ID NO: 4 and SEQ ID NO: 5, and any amplicon thatcomprises SEQ ID NO: 2 or at least 51 consecutive nucleotides of SEQ IDNO: 8 is an aspect of the invention.

The invention provides methods, compositions, and kits useful fordetecting the presence of DNA derived from soybean event MON87712.Certain methods comprise (a) contacting a sample comprising DNA with aprimer set that when used in a nucleic acid amplification reaction withgenomic DNA from a soybean comprising event MON87712 produces anamplicon diagnostic for the event; (b) performing a nucleic acidamplification reaction thereby producing the amplicon; and (c) detectingthe amplicon, wherein said amplicon comprises SEQ ID NO: 1 and/or SEQ IDNO: 2, or at least 51 consecutive nucleotides of SEQ ID NO: 7 or atleast 51 consecutive nucleotides of SEQ ID NO: 8. The invention alsoprovides a method for detection of the event by (a) contacting a samplecomprising DNA with a probe that when used in a hybridization reactionwith genomic DNA from the event hybridizes to a DNA molecule specificfor the event; (b) subjecting the sample and probe to stringenthybridization conditions; and (c) detecting the hybridization of theprobe to the DNA molecule. Kits comprising the methods and compositionsof the invention useful for detecting the presence of DNA derived fromthe event are also provided.

Another aspect of the invention is a method of determining the zygosityof a soybean plant genome comprising soybean event MON87712 DNA in asample comprising: a) contacting the sample with three different primersthat i) when used together in a nucleic acid amplification reaction withsoybean event MON87712 DNA, produces a first amplicon that is diagnosticfor soybean event MON87712; and ii) when used together in a nucleic acidamplification reaction with soybean genomic DNA other than MON87712 DNA,produces a second amplicon that is diagnostic for soybean wild typegenomic DNA other than event MON87712 DNA; b) performing a nucleic acidamplification reaction; and c) detecting said first amplicon and saidsecond amplicon, wherein the presence of said first and second ampliconsis diagnostic of a heterozygous genome in said sample, and wherein thepresence of only said first amplicon is diagnostic of a homozygousgenome in said sample. In one embodiment the primers comprise SEQ ID NO:10, SEQ ID NO: 11 or 15, and SEQ ID NO: 13.

In one embodiment the method of determining the zygosity of a soybeancomprising event MON87712, comprises (a) contacting the samplecomprising soybean DNA with the primer set SEQ ID NO: 10, SEQ ID NO: 11or SEQ ID NO: 15, SEQ ID NO: 13, and the probe set 6FAM™-labeled SEQ IDNO: 12 and VIC™-labeled SEQ ID NO: 14 that when used in a nucleic acidamplification reaction with genomic DNA from a soybean comprising eventMON87712, produces a first amplicon, releasing a fluorescent signal fromthe combination of primers SEQ ID NO: 10 and SEQ ID NO: 11 or 15 and the6FAM™-labeled probe (SEQ ID NO: 12) that is diagnostic for soybean eventMON87712; (b) performing a nucleic acid amplification reaction, therebyproducing the first amplicon; and (c) detecting said first amplicon; and(d) contacting the sample comprising soybean DNA with the primer set,SEQ ID NO: 10 and SEQ ID NO: 13, and the VIC™-labeled probe (SEQ ID NO:14) that when used in a nucleic acid amplification reaction with genomicDNA from soybean plants produces a second amplicon, releasing afluorescent signal that is diagnostic of wild-type soybean genomic DNAhomologous to the soybean genomic region of a transgene insertionidentified as soybean event MON87712; (e) performing a nucleic acidamplification reaction, thereby producing the second amplicon; and (f)detecting said second amplicon; and (g) comparing the first and thesecond amplicons in a sample, wherein the presence of both ampliconsindicates the sample is heterozygous for the transgene insertion.

The invention further provides a method of producing a soybean plantwith increased yield comprising: (a) selfing a MON87712 comprisingsoybean plant, thereby producing a seed; (b) growing said seed toproduce a plurality of progeny plant; and (c) selecting a progeny plantthat comprises MON87712 or a progeny plant with increased yield. Anotheraspect of the invention provides a method of producing a soybean plantwith increased yield comprising: (a) crossing a MON87712 comprisingsoybean plant with a second soybean plant, thereby producing a seed; (b)growing said seed to produce a plurality of progeny plants; and (c)selecting a progeny plant that comprises MON87712 or a progeny plantwith increased yield.

In yet another aspect, the invention provides a method of increasingyield in a crop comprising a) planting a crop plant or seed comprisingevent MON87712; and b) growing said crop plant or seed.

The foregoing and other aspects of the invention will become moreapparent from the following detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Diagrammatical representation of the transgenic insert in thegenome of a soybean comprising event MON87712; [A] corresponds to therelative positions of SEQ ID NO: 1 and SEQ ID NO: 7, both forming thejunction between the 5′ portion of the transgenic insert and the 3′portion of the flanking genomic DNA; [B] corresponds to the relativepositions of SEQ ID NO: 2 and SEQ ID NO: 8, both forming the junctionbetween the 3′ portion of the transgenic insert and the 5′ portion ofthe flanking genomic DNA; [C] corresponds to the relative position ofSEQ ID NO: 3, which contains the soybean genomic flanking region and aportion of the arbitrarily designated 5′ end of the transgenic DNAinsert; [D] corresponds to the relative position of SEQ ID NO: 4, whichcontains the soybean genome flanking region and a portion of thearbitrarily designated 3′ end of the transgenic DNA insert; [E]represents SEQ ID NO: 5, which is the sequence of the transgenic DNAinsert including the BBX32 expression cassette integrated into thegenome of a soybean plant comprising event MON87712; [F] represents SEQID NO: 6, which is the contiguous sequence comprising the 5′ flankinggenomic sequence, the transgenic insert and the 3′ flanking genomicsequence, comprising as represented in the figure from left to right,SEQ ID NO: 3, SEQ ID NO: 5 and SEQ ID NO: 4, in which SEQ ID NOs: 1 and7, and SEQ ID NOs: 2 and 8 are incorporated as set forth above, as thesesequences are present in the genome of a plant comprising eventMON87712; [G] and [H] represent forward primers (SEQ ID NO: 11 or SEQ IDNO: 15, and SEQ ID NO: 13) for event-specific zygosity endpoint TAQMAN®PCR for identification of MON87712 and wild-type allele, respectively;[I] represents reverse primer (SEQ ID NO: 10) for identification ofMON87712 and wild-type alleles; [J] and [K] represent probes (SEQ ID NO:12 and SEQ ID NO: 14) used for event-specific zygosity endpoint TAQMAN®PCR for identification of MON87712 and wild-type allele, respectively.Arrows indicate the direction of 5′ to 3′. LB: refers to the left borderof T-DNA; RB: refers to the right border of T-DNA.

FIG. 2. Meta-analysis of yield data for 9 transgenic events from 4 fieldseasons of field testing in the United States or South America, showingevent 1 comprising MON87712 DNA with the highest yield. Panel A: averageyield increases of transgenic events over their corresponding negativeisolines (bushel/acre). Panel B: average yield increases of transgenicevents over wild type control (bushel/acre). The numbers above each barrepresent % increase.

BRIEF DESCRIPTION OF THE SEQUENCES

SEQ ID NO: 1—A 22 bp nucleotide sequence representing the left borderjunction between the soybean genomic DNA and the integrated DNA insert.This sequence corresponds to positions 3495 to 3516 of SEQ ID NO: 6, andto positions 3495 through 3516 of SEQ ID NO: 3 ([C] of FIG. 1). Inaddition, SEQ ID NO: 1 corresponds to the integrated left border of theexpression cassette at positions 1 through 11 of SEQ ID NO: 5 ([E] ofFIG. 1).

SEQ ID NO: 2—A 22 bp nucleotide sequence representing the right borderjunction between the integrated DNA insert and the soybean genomic DNA.This sequence corresponds to positions 5509 to 5530 of SEQ ID NO: 6, andto positions 90 through 111 of SEQ ID NO: 4 ([D] of FIG. 1). Inaddition, SEQ ID NO: 2 corresponds to positions 2004 through 2014 SEQ IDNO: 5 ([E] of FIG. 1).

SEQ ID NO: 3—A 3605 bp nucleotide sequence including the 5′ soybeangenomic sequence (3505 bp) flanking the inserted DNA of event MON87712plus a region (100 bp) of the integrated DNA. This sequence correspondsto positions 1 through 3605 of SEQ ID NO: 6. It is believed that thesequence at position 3451 to 3463 of SEQ ID NO: 3 may contain anywherefrom 13 to 100 individual c residues in a row.

SEQ ID NO: 4—A 2065 bp nucleotide sequence including the 3′ soybeangenomic sequence (1965 bp) flanking the inserted DNA of event MON87712plus a region (100 bp) of the integrated DNA. This sequence correspondsto positions 5420 through 7484 of SEQ ID NO: 6.

SEQ ID NO: 5—The sequence of the integrated expression cassetteimparting increased yield, including the left and the right bordersequences after integration. SEQ ID NO: 5 corresponds to nucleotidepositions 3506 through 5519 of SEQ ID NO: 6.

SEQ ID NO: 6—A 7484 bp nucleotide sequence representing the contig ofthe 5′ sequence flanking the inserted DNA of MON87712 (SEQ ID NO: 3),the sequence of the integrated DNA insert (SEQ ID NO: 5) and the 3′sequence flanking the inserted DNA of MON87712 (SEQ ID NO: 4). It isbelieved that the sequence at position 3451 to 3463 of SEQ ID NO: 6 maycontain anywhere from 13 to 100 individual c residues in a row.

SEQ ID NO: 7: A 100 bp nucleotide sequence representing the left borderjunction between soybean genomic DNA and the integrated DNA insert. Thissequence corresponds to positions 3456 to 3555 of SEQ ID NO: 6, and topositions 3456 through 3555 of SEQ ID NO: 3 ([C] of FIG. 1). Inaddition, SEQ ID NO: 7 corresponds to the integrated left border of theexpression cassette at positions 1 through 50 of SEQ ID NO: 5 ([E] ofFIG. 1).

SEQ ID NO: 8—A 100 bp nucleotide sequence representing the right borderjunction between the integrated DNA insert and the soybean genomic DNA.This sequence corresponds to positions 5470 to 5569 of SEQ ID NO: 6, andto positions 51 through 150 of SEQ ID NO: 4 ([D] of FIG. 1). Inaddition, SEQ ID NO: 8 corresponds to positions 1965 through 2014 SEQ IDNO: 5 ([E] of FIG. 1).

SEQ ID NO: 9—The nucleotide sequence of the BBX32 expression cassette ofpMON83132.

SEQ ID NO: 10—The reverse primer sequence used in event-specificzygosity endpoint TAQMAN PCR assay for identification of both MON87712DNA/allele and wild-type DNA/allele. SEQ ID NO: 10 corresponds tonucleotide positions 5517 to 5546 of SEQ ID NO: 6. Production of a PCRamplicon using the combination of primers SEQ ID NO: 10 and SEQ ID NO:11 or 15 is a positive result for the presence of event MON87712.

SEQ ID NO: 11—The forward primer sequence used to identify MON87712event and the zygosity of MON87712 event. SEQ ID NO: 11 corresponds tonucleotide positions 5435 to 5458 of SEQ ID NO: 6.

SEQ ID NO: 12—The probe sequence used to identify MON87712 event and thezygosity of MON87712 event. SEQ ID NO: 12 is a 6FAM™-labeled syntheticoligonucleotide. Release of a fluorescent signal in an amplificationreaction using primers SEQ ID NO: 10 and SEQ ID NO: 11 or 15 incombination with the 6FAM™-labeled probe in a TAQMAN® assay isdiagnostic of event MON87712.

SEQ ID NO: 13—The forward primer sequence used in event-specificzygosity endpoint TAQMAN® PCR for identification of wild-typeDNA/allele. Production of a PCR amplicon using primers SEQ ID NO: 13 andSEQ ID NO: 10 is diagnostic of wild-type DNA.

SEQ ID NO: 14—The probe sequence used to determine the presence ofsoybean wild-type DNA/allele. SEQ ID NO: 14 is a VIC™-labeled syntheticoligonucleotide. Release of a fluorescent signal in an amplificationreaction using primers SEQ ID NO: 10 and SEQ ID NO: 13 in combinationwith the VIC™-labeled probe in a TAQMAN® assay is diagnostic of thewild-type allele in a zygosity assay.

SEQ ID NO: 15—An alternative forward primer sequence used to identifyMON87712 event and the zygosity of MON87712 event. SEQ ID NO: 15corresponds to nucleotide positions 5432 to 5455 of SEQ ID NO: 6.

SEQ ID NO: 16—The sequence of primer SQ3983 used to identify MON87712event. Production of a 97 bp PCR amplicon using the combination ofprimers SQ3983 and SQ22982 (SEQ ID NO: 17) is a positive result for thepresence of event MON87712.

SEQ ID NO: 17—The sequence of primer SQ22982 used to identify MON87712event.

SEQ ID NO: 18—The sequence of probe PB10453 used to identify MON87712event. It is a 6FAM™-labeled synthetic oligonucleotide.

SEQ ID NO: 19—The sequence of primer SQ1532 used as an internal controlin end-point TAQMAN® assays.

SEQ ID NO: 20—The sequence of primer SQ1533 used as an internal controlin end-point TAQMAN® assays.

SEQ ID NO: 21—The sequence of a VIC™-labeled synthetic oligonucleotideprobe PB0359 used as an internal control in end-point TAQMAN® assays.

DETAILED DESCRIPTION

The following definitions and methods are provided to better define thepresent invention and to guide those of ordinary skill in the art in thepractice of the present invention. Unless otherwise noted, terms are tobe understood according to conventional usage by those of ordinary skillin the relevant art. Definitions of common terms in molecular biologymay also be found in Rieger et al., Glossary of Genetics: Classical andMolecular, 5th edition, Springer-Verlag: New York, 1991; and Lewin,Genes V, Oxford University Press: New York, 1994.

The present invention provides transgenic soybean event MON87712. Theterm “event” as used herein refers to DNA molecules produced as a resultof inserting transgenic DNA into a plant's genome at a particularlocation on a chromosome. Event MON87712 refers to the DNA moleculesproduced as a result of the insertion of transgenic DNA having asequence provided herein as SEQ ID NO: 5 into a particular chromosomallocation in the Glycine max genome. Plants, seeds, progeny, cells, andplant parts thereof comprising event MON87712 are also provided in thepresent invention. A sample of seed comprising MON87712 has beendeposited with American Type Culture Collection (ATCC) under AccessionNo. PTA-10296 on Aug. 20, 2009. Plants comprising MON87712 exhibitincreased yield.

As used herein, the term “soybean” means Glycine max and includes allplant varieties that can be bred with soybean, including wild soybeanspecies as well as those plants belonging to Glycine soja that permitbreeding between species.

A transgenic “event” is produced by transformation of plant cells withheterologous DNA, i.e., a nucleic acid construct that comprises atransgene of interest, regeneration of a population of independentlytransformed transgenic plants resulting from the insertion of thetransgene into the genome of the plant, and selection of a particularplant with desirable molecular characteristics, such as insertion of asingle copy of the transgene into a particular genome location,integrity of the transgenic DNA, and an enhanced trait such as increasedyield. A plant comprising the event can refer to the originaltransformant that includes the transgene inserted into the particularlocation in the plant's genome. A plant comprising the event can alsorefer to progeny of the original transformant that retain the transgeneat the same particular location in the plant's genome. Such progeny maybe produced by selfing or by a sexual outcross between the transformant,or its progeny, and another plant. Such another plant may be atransgenic plant comprising the same or a different transgene; or may bea non-transgenic plant, such as one from a different variety. Theresulting progeny may be homozygous or heterozygous for event MON87712DNA (inserted DNA and flanking DNA). Even after repeated back-crossingto a recurrent parent, the event DNA from the transformed parent ispresent in the progeny of the cross at the same genomic location.

A DNA molecule comprising event MON87712 refers to a DNA moleculecomprising at least a portion of the inserted transgenic DNA (providedas SEQ ID NO: 5) and at least a portion of the flanking genomic DNAimmediately adjacent to the inserted DNA. As such, a DNA moleculecomprising event MON87712 has a nucleotide sequence representing atleast a portion of the transgenic DNA insert and at least a portion ofthe particular region of the genome of the plant into which thetransgenic DNA was inserted. The arrangement of the inserted DNA inevent MON87712 in relation to the surrounding plant genome is specificand unique to event MON87712 and as such the nucleotide sequence of sucha DNA molecule is diagnostic and identifying for event MON87712.Examples of the sequence of such a DNA molecule are provided herein asSEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 7,SEQ ID NO: 8 and SEQ ID NO: 6. Such a DNA molecule is also an integralpart of the chromosome of a plant that comprises event MON87712 and maybe passed on to progenies of the plant.

As used herein, a “recombinant DNA molecule” is a DNA moleculecomprising a combination of DNA molecules that would not naturally occurtogether and is the result of human intervention, e.g., a DNA moleculethat is comprised of a combination of at least two DNA moleculesheterologous to each other, and/or a DNA molecule that is artificiallysynthesized and comprises a polynucleotide sequence that deviates fromthe polynucleotide sequence that would normally exist in nature, and/ora DNA molecule that comprises a transgene artificially incorporated intoa host cell's genomic DNA and the associated flanking DNA of the hostcell's genome. An example of a recombinant DNA molecule is a DNAmolecule described herein resulting from the insertion of the transgeneinto the Glycine max genome, which may ultimately result in theexpression of a recombinant RNA and/or protein molecule in thatorganism. The nucleotide sequence or any fragment derived therefromwould also be considered a recombinant DNA molecule if the DNA moleculecan be extracted from cells, or tissues, or homogenate from a plant orseed or plant tissue; or can be produced as an amplicon from extractedDNA or RNA from cells, or tissues, or homogenate from a plant or seed orplant tissue, any of which is derived from such materials derived from aplant comprising event MON87712. For that matter, the junction sequencesas set forth at SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 7 and SEQ ID NO:8, and nucleotide sequences derived from event MON87712 that alsocontain these junction sequences are considered to be recombinant DNA,whether these sequences are present within the genome of the cellscomprising event MON87712 or present in detectable amounts in tissues,progeny, biological samples or commodity products derived from plantscomprising event MON87712. As used herein, the term “transgene” refersto a polynucleotide molecule artificially incorporated into a hostcell's genome. Such transgene may be heterologous to the host cell. Theterm “transgenic plant” refers to a plant comprising such a transgene. A“transgenic plant” includes a plant, plant part, a plant cell or seedwhose genome has been altered by the stable integration of recombinantDNA. A transgenic plant includes a plant regenerated from anoriginally-transformed plant cell and progeny transgenic plants fromlater generations or crosses of a transformed plant. As a result of suchgenomic alteration, the transgenic plant is distinctly different fromthe related wild type plant. An example of a transgenic plant is a plantdescribed herein as comprising event MON87712.

As used herein, the term “heterologous” refers to a sequence which isnot normally present in a given host genome in the genetic context inwhich the sequence is currently found. In this respect, the sequence maybe native to the host genome, but be rearranged with respect to othergenetic sequences within the host sequence.

The present invention provides DNA molecules and their correspondingnucleotide sequences. As used herein, the terms “DNA sequence”,“nucleotide sequence” and “polynucleotide sequence” refer to thesequence of nucleotides of a DNA molecule, usually presented from the 5′(upstream) end to the 3′ (downstream) end. The nomenclature used hereinis that required by Title 37 of the United States Code of FederalRegulations § 1.822 and set forth in the tables in WIPO Standard ST.25(1998), Appendix 2, Tables 1 and 3. The present invention is disclosedwith reference to only one strand of the two nucleotide sequence strandsthat are provided in transgenic event MON87712. Therefore, byimplication and derivation, the complementary sequences, also referredto in the art as the complete complement or the reverse complementarysequences, are within the scope of the present invention and aretherefore also intended to be within the scope of the subject matterclaimed.

The nucleotide sequence corresponding to the complete nucleotidesequence of the inserted transgenic DNA and substantial segments of theGlycine max genomic DNA flanking either end of the inserted transgenicDNA is provided herein as SEQ ID NO: 6. A subsection of this is theinserted transgenic DNA provided as SEQ ID NO: 5. The nucleotidesequence of the genomic DNA flanking the 5′ end of the insertedtransgenic DNA and a portion of the 5′ end of the inserted DNA isprovided herein as SEQ ID NO: 3. The nucleotide sequence of the genomicDNA flanking the 3′ end of the inserted transgenic DNA and a portion ofthe 3′ end of the inserted DNA is provided herein as SEQ ID NO: 4. Theregion spanning the location where the transgenic DNA connects to and islinked to the genomic DNA is referred to herein as the junction. A“junction sequence” or “junction region” refers to a DNA sequence and/orcorresponding DNA molecule that spans the inserted transgenic DNA andthe adjacent flanking genomic DNA. Examples of a junction sequence ofevent MON87712 are provided herein as SEQ ID NO: 1, SEQ ID NO: 2, SEQ IDNO: 7, and SEQ ID NO: 8. The identification of one of these junctionsequences in a nucleotide molecule derived from a soybean plant or seedis conclusive that the DNA was obtained from event MON87712 and isdiagnostic for the presence of DNA from event MON87712. SEQ ID NO: 1 isa 22 bp nucleotide sequence spanning the junction between the genomicDNA and the 5′ end of the inserted DNA. SEQ ID NO: 7 is a 100 bpnucleotide sequence spanning the junction between the genomic DNA andthe 5′ end of the inserted DNA. SEQ ID NO: 2 is a 22 bp nucleotidesequence spanning the junction between the genomic DNA and the 3′ end ofthe inserted DNA. SEQ ID NO: 8 is a 100 bp nucleotide sequence spanningthe junction between the genomic DNA and the 3′ end of the inserted DNA.Any segment of DNA derived from transgenic event MON87712 that includesthe consecutive nucleotides of SEQ ID NO: 1 or 51 consecutivenucleotides, 55 consecutive nucleotides, 60 consecutive nucleotides, 65consecutive nucleotides, 70 consecutive nucleotides, 75 consecutivenucleotides, 80 consecutive nucleotides, 85 consecutive nucleotides, 90consecutive nucleotides, 95 consecutive nucleotides, or all of thenucleotides of SEQ ID NO: 7 is within the scope of the presentinvention. Any segment of DNA derived from transgenic event MON87712that includes the consecutive nucleotides of SEQ ID NO: 2 or 51consecutive nucleotides, 55 consecutive nucleotides, 60 consecutivenucleotides, 65 consecutive nucleotides, 70 consecutive nucleotides, 75consecutive nucleotides, 80 consecutive nucleotides, 85 consecutivenucleotides, 90 consecutive nucleotides, 95 consecutive nucleotides, orall of the nucleotides of SEQ ID NO: 8 is within the scope of thepresent invention. In addition, any polynucleotide molecule comprising asequence complementary to any of the sequences described within thisparagraph is within the scope of the present invention. FIG. 1 is anillustration of the transgenic DNA insert in the genome of a soybeancomprising event MON87712, and the relative positions of SEQ ID NOs: 1-8arranged 5′ to 3′. The present invention also provides a nucleic acidmolecule comprising a polynucleotide having a sequence that is at least90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to thefull-length of SEQ ID NO: 6.

The present invention further provides exemplary DNA molecules that canbe used either as primers or probes for diagnosing the presence of DNAderived from event MON87712 in a sample. Such primers or probes arespecific for a target nucleic acid sequence and as such are useful forthe identification of event MON87712 nucleic acid sequence by themethods of the invention described herein.

A “probe” is an isolated nucleic acid to which is attached a detectablelabel or reporter molecule, e.g., a radioactive isotope, ligand,chemiluminescent agent, or enzyme. Such a probe is complementary to astrand of a target nucleic acid, in the case of the present invention,to a strand of genomic DNA from a soybean comprising event MON87712whether from a soybean plant or from a sample that comprises DNA fromthe event. Probes according to the present invention include not onlydeoxyribonucleic or ribonucleic acids but also polyamides and otherprobe materials that bind specifically to a target DNA sequence and thedetection of such binding can be used to diagnose/determine/confirm thepresence of that target DNA sequence in a particular sample.

A “primer” is typically an isolated polynucleotide that is designed foruse in specific annealing or hybridization methods to hybridize to acomplementary target DNA strand to form a hybrid between the primer andthe target DNA strand, and then extended along the target DNA strand bya polymerase, e.g., a DNA polymerase. A pair of primers may be used withtemplate DNA, such as a sample of Glycine max genomic DNA, in a thermalor isothermal amplification, such as polymerase chain reaction (PCR), orother nucleic acid amplification methods, to produce an amplicon, wherethe amplicon produced from such reaction would have a DNA sequencecorresponding to sequence of the template DNA located between the twosites where the primers hybridized to the template. As used herein, an“amplicon” is a piece or fragment of DNA that has been synthesized usingamplification techniques, i.e. the product of an amplification reaction.In one embodiment of the invention, an amplicon diagnostic for eventMON87712 comprises a sequence not naturally found in the Glycine maxgenome. Primer pairs, as used in the present invention, are intended torefer to use of two primers binding opposite strands of a doublestranded nucleotide segment for the purpose of amplifying linearly thepolynucleotide segment between the positions targeted for binding by theindividual members of the primer pair, typically in a thermal orisothermal amplification reaction or other nucleic acid amplificationmethods. Exemplary DNA molecules useful as primers are provided as SEQID NOs: 10-11, SEQ ID NO: 13 and SEQ ID NO: 15. The primer pair providedas SEQ ID NO: 10 and SEQ ID NO: 11 or 15 may be used as a first DNAmolecule and a second DNA molecule that is different from the first DNAmolecule, and both molecules are each of sufficient length ofconsecutive nucleotides of either SEQ ID NO: 4, SEQ ID NO: 5, or SEQ IDNO: 6 or the complements thereof to function as DNA primers so that,when used together in an amplification reaction with template DNAderived from event MON87712, an amplicon that is specific and unique totransgenic event MON87712 would be produced. The use of the term“amplicon” specifically excludes primer-dimers that may be formed in theDNA amplification reaction.

Probes and primers according to the present invention may have completesequence identity to the target sequence, although primers and probesdiffering from the target sequence that retain the ability to hybridizepreferentially to target sequences may be designed by conventionalmethods. In order for a nucleic acid molecule to serve as a primer orprobe it needs only be sufficiently complementary in sequence to be ableto form a stable double-stranded structure under the particular solventand salt concentrations employed. Any nucleic acid hybridization oramplification method can be used to identify the presence of transgenicDNA from event MON87712 in a sample. Probes and primers are generally atleast about 11 nucleotides, at least about 18 nucleotides, at leastabout 24 nucleotides, and at least about 30 nucleotides or more inlength. Such probes and primers hybridize specifically to a targetsequence under high stringency hybridization conditions.

Methods for preparing and using probes and primers are described, forexample, in Molecular Cloning: A Laboratory Manual, 2nd ed., vol. 1-3,ed. Sambrook et al., Cold Spring Harbor Laboratory Press, Cold SpringHarbor, N.Y., 1989 (hereinafter, “Sambrook et al., 1989”); CurrentProtocols in Molecular Biology, ed. Ausubel et al., Greene Publishingand Wiley-Interscience, New York, 1992 (with periodic updates)(hereinafter, “Ausubel et al., 1992”); and Innis et al., PCR Protocols:A Guide to Methods and Applications, Academic Press: San Diego, 1990.PCR-primer pairs can be derived from a known sequence, for example, byusing computer programs intended for that purpose such as Primer(Version 0.5, © 1991, Whitehead Institute for Biomedical Research,Cambridge, Mass.).

Primers and probes based on the flanking DNA and insert sequencesdisclosed herein can be used to confirm the disclosed sequences by knownmethods, e.g., by re-cloning and sequencing such sequences.

The nucleic acid probes and primers of the present invention hybridizeunder stringent conditions to a target DNA sequence. Any nucleic acidhybridization or amplification method can be used to identify thepresence of DNA from a transgenic event in a sample. Nucleic acidmolecules or fragments thereof are capable of specifically hybridizingto other nucleic acid molecules under certain circumstances. As usedherein, two nucleic acid molecules are said to be capable ofspecifically hybridizing to one another if the two molecules are capableof forming an anti-parallel, double-stranded nucleic acid structure. Anucleic acid molecule is said to be the “complement” of another nucleicacid molecule if they exhibit complete complementarity. As used herein,molecules are said to exhibit “complete complementarity” when everynucleotide of one of the molecules is complementary to a nucleotide ofthe other. Two molecules are said to be “minimally complementary” ifthey can hybridize to one another with sufficient stability to permitthem to remain annealed to one another under at least “low-stringency”conditions. Similarly, the molecules are said to be “complementary” ifthey can hybridize to one another with sufficient stability to permitthem to remain annealed to one another under “high-stringency”conditions. Stringency conditions are described by Sambrook et al.,1989, and by Haymes et al., In: Nucleic Acid Hybridization, A PracticalApproach, IRL Press, Washington, D.C. (1985). Departures from completecomplementarity are therefore permissible, as long as such departures donot completely preclude the capacity of the molecules to form adouble-stranded structure. In order for a nucleic acid molecule to serveas a primer or probe it need only be sufficiently complementary insequence to be able to form a stable double-stranded structure under theparticular solvent and salt concentrations employed.

As used herein, a substantially homologous sequence is a nucleic acidsequence that will specifically hybridize to the complement of thenucleic acid sequence to which it is being compared under highstringency conditions. Appropriate stringency conditions that promoteDNA hybridization, for example, 6.0× sodium chloride/sodium citrate(SSC) at about 45° C., followed by a wash of 2.0×SSC at 50° C., areknown to those skilled in the art or can be found in Current Protocolsin Molecular Biology, John Wiley & Sons, N.Y. (1989), 6.3.1-6.3.6. Forexample, the salt concentration in the wash step can be selected from alow stringency of about 2.0×SSC at 50° C. to a high stringency of about0.2×SSC at 50° C. In addition, the temperature in the wash step can beincreased from low stringency conditions at room temperature, about 22°C., to high stringency conditions at about 65° C. Both temperature andsalt may be varied, or either the temperature or the salt concentrationmay be held constant while the other variable is changed. In oneembodiment, a nucleic acid of the present invention will specificallyhybridize to one or more of the nucleic acid molecules set forth in SEQID NO: 1, SEQ ID NO: 2, SEQ ID NO: 7 and SEQ ID NO: 8, or complements orfragments thereof under high stringency conditions. The hybridization ofthe probe to the target DNA molecule can be detected by any number ofmethods known to those skilled in the art. These can include, but arenot limited to, fluorescent tags, radioactive tags, antibody based tags,and chemiluminescent tags.

Regarding the amplification of a target nucleic acid sequence (e.g., byPCR) using a particular amplification primer pair, “stringentconditions” are conditions that permit the primer pair to hybridize onlyto the target nucleic acid sequence to which a primer having thecorresponding wild-type sequence (or its complement) would bind andpreferably to produce a unique amplification product, the amplicon, in aDNA amplification reaction. Examples of DNA amplification methodsinclude PCR, Recombinase Polymerase Amplification (RPA) (see for exampleU.S. Pat No. 7,485,428), Strand Displacement Amplification (SDA) (seefor example, U.S. Pat. Nos. 5,455,166 and 5,470,723),Transcription-Mediated Amplification (TMA) (see for example, Guatelli etal., Proc. Natl. Acad. Sci. USA 87:1874-1878 (1990)), Rolling CircleAmplification (RCA) (see for example, Fire and Xu, Proc. Natl. Acad Sci.USA 92:4641-4645 (1995); Lui, et al., J. Am. Chem. Soc. 118:1587-1594(1996); Lizardi, et al., Nature Genetics 19:225-232 (1998), U.S. Pat.Nos. 5,714,320 and 6,235,502)), Helicase Dependant Amplification (HDA)(see for example Vincent et al., EMBO Reports 5 (8): 795-800 (2004);U.S. Pat. No. 7,282,328), and Multiple Displacement Amplification (MDA)(see for example Dean et. al., Proc. Natl. Acad Sci. USA 99:5261-5266(2002)).

The term “specific for (a target sequence)” indicates that a probe orprimer hybridizes under stringent hybridization conditions only to thetarget sequence in a sample comprising the target sequence.

As used herein, the term “isolated” refers to at least partiallyseparating a molecule from other molecules normally associated with itin its native or natural state. In one embodiment, the term “isolated”refers to a DNA molecule that is at least partially separated from thenucleic acids that normally flank the DNA molecule in its native ornatural state. Thus, DNA molecules fused to regulatory or codingsequences with which they are not normally associated, for example asthe result of recombinant techniques, are considered isolated herein.Such molecules are considered isolated even when integrated into thechromosome of a host cell or present in a nucleic acid solution withother DNA molecules.

Any number of methods well known to those skilled in the art can be usedto isolate and manipulate a DNA molecule, or fragment thereof, disclosedin the present invention. For example, PCR (polymerase chain reaction)technology can be used to amplify a particular starting DNA moleculeand/or to produce variants of the original molecule. DNA molecules, orfragments thereof, can also be obtained by other techniques such as bydirectly synthesizing the fragment by chemical means, as is commonlypracticed by using an automated oligonucleotide synthesizer.

It would be advantageous to be able to detect the presence oftransgene/genomic DNA of a particular plant in order to determinewhether progeny of a sexual cross contain the transgene/genomic DNA ofinterest. In addition, a method for detecting a particular plant wouldbe helpful when complying with regulations requiring the pre-marketapproval and labeling of foods derived from the transgenic crop plants.

The presence of a transgene may be detected by any well known nucleicacid detection method such as the polymerase chain reaction (PCR) or DNAhybridization using nucleic acid probes. These detection methodsgenerally focus on frequently used genetic elements, such as promoters,terminators, marker genes, etc. As a result, such methods may not beuseful for discriminating between different transformation events,particularly those produced using the same DNA construct unless thesequence of chromosomal DNA adjacent to the inserted DNA (“flankingDNA”) is known. An event-specific PCR assay is discussed, for example,by Taverniers et al. (J. Agric. Food Chem., 53: 3041-3052, 2005) inwhich an event-specific tracing system for transgenic maize lines Bt11,Bt176, and GA21 and for canola event GT73 was demonstrated. In thisstudy, event-specific primers and probes were designed based upon thesequences of the genome/transgene junctions for each event. Transgenicplant event specific DNA detection methods have also been described inU.S. Pat. Nos. 6,893,826; 6,825,400; 6,740,488; 6,733,974; 6,689,880;6,900,014 and 6,818,807.

The DNA molecules and corresponding nucleotide sequences provided hereinare therefore useful for, among other things, identifying eventMON87712, selecting plant varieties or hybrids comprising eventMON87712, detecting the presence of DNA derived from event MON87712 in asample, and monitoring samples for the presence and/or absence of eventMON87712 or plants and plant parts comprising event MON87712.

The present invention provides plants, progeny, seeds, plant cells,plant parts (such as pollen, ovule, pod, flower, root or stem tissue,fibers, and leaf), and commodity products. These plants, progeny, seeds,plant cells, plant parts, and commodity products contain a detectableamount of a polynucleotide of the present invention, i.e., such as apolynucleotide comprising at least one of the sequences provided as theconsecutive nucleotides of SEQ ID NO: 1, the consecutive nucleotides ofSEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7, or atleast 51 consecutive nucleotides of SEQ ID NO: 8. Plants, progeny,seeds, plant cells, plant parts and commodity products of the presentinvention may also contain one or more additional transgenes. Suchtransgene may be any nucleotide sequence encoding a protein or RNAmolecule conferring a desirable trait including but not limited toincreased insect resistance, increased water use efficiency or droughttolerance, increased yield performance, increased nitrogen useefficiency or increase tolerance to nitrogen stress such as high or lownitrogen supply, increased seed quality, increased disease resistance,improved nutritional quality, and/or increased herbicide tolerance, suchas glyphosate or dicamba tolerance, in which the desirable trait ismeasured with respect to a comparable plant lacking such additionaltransgene.

The present invention provides plants, progeny, seeds, plant cells, andplant part such as pollen, ovule, pod, flower, root or stem tissue, andleaf derived from a transgenic plant comprising event MON87712. Arepresentative sample of seed comprising event MON87712 has beendeposited according to the Budapest Treaty for the purpose of enablingthe present invention. The repository selected for receiving the depositis the American Type Culture Collection (ATCC) having an address at10801 University Boulevard, Manassas, Va. USA, Zip Code 20110. The ATCCrepository has assigned the accession No. PTA-10296 to event MON87712containing seed.

The present invention provides a microorganism comprising a DNA moleculehaving a nucleotide sequence selected from the group consisting of theconsecutive nucleotides of SEQ ID NO: 1, the consecutive nucleotides ofSEQ ID NO: 2, or at least 51 consecutive nucleotides of SEQ ID NO: 7, orof SEQ ID NO: 8 present in its genome. An example of such amicroorganism is a transgenic plant cell. Microorganisms, such as aplant cell of the present invention, are useful in many industrialapplications, including but not limited to: (i) use as research tool forscientific inquiry or industrial research; (ii) use in culture forproducing endogenous or recombinant carbohydrate, lipid, nucleic acid,enzymes or protein products or small molecules that may be used forsubsequent scientific research or as industrial products; and (iii) usewith modern plant tissue culture techniques to produce transgenic plantsor plant tissue cultures that may then be used for agricultural researchor production. The production and use of microorganisms such astransgenic plant cells utilizes modern microbiological techniques andhuman intervention to produce a man-made, unique microorganism. In thisprocess, recombinant DNA is inserted into a plant cell's genome tocreate a transgenic plant cell that is separate and unique fromnaturally occurring plant cells. This transgenic plant cell can then becultured much like bacteria and yeast cells using modern microbiologytechniques and may exist in an undifferentiated, unicellular state. Thenew plant cell's genetic composition and phenotype is a technical effectcreated by the integration of the heterologous DNA into the genome ofthe cell. Another aspect of the present invention is a method of using amicroorganism of the present invention. Methods of using microorganismsof the present invention, such as transgenic plant cells, include (i)methods of producing transgenic cells by integrating recombinant DNAinto genome of the cell and then using this cell to derive additionalcells possessing the same heterologous DNA; (ii) methods of culturingcells that contain recombinant DNA using modern microbiology techniques;(iii) methods of producing and purifying endogenous or recombinantcarbohydrate, lipid, nucleic acid, enzymes or protein products fromcultured cells; and (iv) methods of using modern plant tissue culturetechniques with transgenic plant cells to produce transgenic plants ortransgenic plant tissue cultures.

As used herein, “progeny” includes any plant, seed, plant cell, and/orregenerable plant part comprising the event DNA derived from an ancestorplant and/or a polynucleotide having at least one of the sequencesprovided as the consecutive nucleotides of SEQ ID NO: 1, the consecutivenucleotides of SEQ ID NO: 2, or at least 51 consecutive nucleotides ofSEQ ID NO: 7, or at least 51 consecutive nucleotides of SEQ ID NO: 8.Plants, progeny, and seeds may be homozygous or heterozygous for thetransgene. Progeny may be grown from seeds produced by a plantcomprising event MON87712 and/or from seeds produced by a plantfertilized with pollen from a plant comprising event MON87712.

Progeny plants may be self-pollinated (also known as “selfing”) togenerate a true breeding line of plants, i.e., plants homozygous for thetransgene. Alternatively, progeny plants may be outcrossed, e.g., bredwith another plant, to produce a varietal or a hybrid seed or plant. Theother plant may be transgenic or nontransgenic. A varietal or hybridseed or plant of the present invention may thus be derived by crossing afirst parent that lacks the specific and unique DNA of event MON87712with a second parent comprising event MON87712, resulting in a hybridcomprising the specific and unique DNA of event MON87712. Each parentcan be a hybrid or an inbred/variety, so long as the cross or breedingresults in a plant or seed of the present invention, i.e., a seed havingat least one allele comprising the specific and unique DNA of eventMON87712 and/or the consecutive nucleotides of SEQ ID NO: 1 or SEQ IDNO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7 or at least51 consecutive nucleotides of SEQ ID NO: 8. Two different transgenicplants may thus be mated to produce hybrid offspring that contain twoindependently segregating, added, exogenous genes. For example, a plantcomprising event MON87712 with increased yield can be crossed withanother transgenic plant, such as one tolerant to glyphosate, to producea plant having the characteristics of both transgenic parents. Selfingof appropriate progeny can produce plants that are homozygous for bothadded, exogenous genes. Back-crossing to a parental plant andout-crossing with a non-transgenic plant are also contemplated, as isvegetative propagation. Descriptions of other breeding methods that arecommonly used for different traits and crops can be found in one ofseveral references, e.g., Fehr, in Breeding Methods for CultivarDevelopment, Wilcox J. ed., American Society of Agronomy, Madison Wis.(1987). Sexually crossing one plant with another plant, i.e.,cross-pollinating, may be accomplished or facilitated by humanintervention, for example: by human hands collecting the pollen of oneplant and contacting this pollen with the style or stigma of a secondplant; by human hands and/or human actions removing, destroying, orcovering the stamen or anthers of a plant (e.g., by manual interventionor by application of a chemical gametocide) so that naturalself-pollination is prevented and cross-pollination would have to takeplace in order for fertilization to occur; by human placement ofpollinating insects in a position for “directed pollination” (e.g., byplacing beehives in orchards or fields or by caging plants withpollinating insects); by human opening or removing of parts of theflower to allow for placement or contact of foreign pollen on the styleor stigma; by selective placement of plants (e.g., intentionallyplanting plants in pollinating proximity); and/or by application ofchemicals to precipitate flowering or to foster receptivity (of thestigma for pollen).

In practicing this method, the step of sexually crossing one plant withitself, i.e., self-pollinating or selfing, may be accomplished orfacilitated by human intervention, for example: by human handscollecting the pollen of the plant and contacting this pollen with thestyle or stigma of the same plant and then optionally preventing furtherfertilization of the plant; by human hands and/or actions removing,destroying, or covering the stamen or anthers of other nearby plants(e.g., by detasseling or by application of a chemical gametocide) sothat natural cross-pollination is prevented and self-pollination wouldhave to take place in order for fertilization to occur; by humanplacement of pollinating insects in a position for “directedpollination” (e.g., by caging a plant alone with pollinating insects);by human manipulation of the flower or its parts to allow forself-pollination; by selective placement of plants (e.g., intentionallyplanting plants beyond pollinating proximity); and/or by application ofchemicals to precipitate flowering or to foster receptivity (of thestigma for pollen).

The present invention provides a plant part that is derived from a plantcomprising event MON87712. As used herein, a “plant part” refers to anypart of a plant that is comprised of material derived from a plantcomprising event MON87712. Plant parts include but are not limited tocells, pollen, ovule, pod, flower, root or stem tissue, fibers, andleaf. Plant parts may be viable, nonviable, regenerable, and/ornon-regenerable.

The present invention provides a commodity product that is derived froma plant comprising event MON87712. As used herein, a “commodity product”refers to any composition or product that is comprised of materialderived from a plant, seed, plant cell, or plant part comprising eventMON87712. Commodity products may be sold to consumers and may be viableor nonviable. Nonviable commodity products include but are not limitedto nonviable seeds and grains; processed seeds, seed parts, and plantparts; dehydrated plant tissue, frozen plant tissue, and processed planttissue; seeds and plant parts processed for animal feed for terrestrialand/or aquatic animal consumption, oil, meal, flour, flakes, bran,fiber, and any other food for human consumption; and biomasses and fuelproducts. Processed soybeans are the largest source of protein feed andvegetable oil in the world. Soybeans and soybean oils from plantscomprising MON87712 can be used in the manufacture of many differentproducts, not limited to, nontoxic plastics, printing inks, lubricants,waxes, hydraulic fluids, electric transformer fluids, solvents,cosmetics, and hair care products. Soybeans and oils of plantscomprising MON87712 can be suitable for use in a variety of soy foodsmade from whole soybeans, such as soymilk, soy nut butter, natto, andtempeh, and soy foods made from processed soybeans and soybean oil,including soybean meal, soy flour, soy protein concentrate, soy proteinisolates, texturized soy protein concentrate, hydrolyzed soy protein,whipped topping, cooking oil, salad oil, shortening, and lecithin. Wholesoybeans are also edible, and are typically sold to consumers raw,roasted, or as edamame. Soymilk, which is typically produced by soakingand grinding whole soybeans, may be consumed without other processing,spray-dried, or processed to form soy yogurt, soy cheese, tofu, or yuba.Viable commodity products include but are not limited to seeds and plantcells. A plant comprising event MON87712 can thus be used to manufactureany commodity product typically acquired from a soybean plant. Any suchcommodity product that is derived from the plants comprising eventMON87712 may contain at least a detectable amount of the specific andunique DNA corresponding to event MON87712, and specifically may containa detectable amount of a polynucleotide having a nucleotide sequence ofthe consecutive nucleotides of SEQ ID NO: 1, the consecutive nucleotidesof SEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7 orat least 51 consecutive nucleotides of SEQ ID NO: 8. Any standard methodof detection for polynucleotide molecules may be used, including methodsof detection disclosed herein. A commodity product is within the scopeof the present invention if there is any detectable amount of theconsecutive nucleotides of SEQ ID NO: 1, the consecutive nucleotides ofSEQ 1D NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7 or atleast 51 consecutive nucleotides of SEQ ID NO: 8 in the commodityproduct.

The plant, progeny, seed, plant cell, plant part (such as pollen, ovule,pod, flower, root or stem tissue, and leaf), and commodity products ofthe present invention are therefore useful for, among other things,growing plants for the purpose of producing seed and/or plant partscomprising event MON87712 for agricultural purposes, producing progenycomprising event MON87712 for plant breeding and research purposes, usewith microbiological techniques for industrial and researchapplications, and sale to consumers.

The present invention provides methods for producing plants withincreased yield and plants comprising event MON87712. An event MON87712containing plant was produced by an Agrobacterium mediatedtransformation method similar to that described in U.S. Pat. No.5,914,451, using an inbred soybean line with the construct pMON83132(Table 1). Construct pMON83132 contains a plant expression cassette forexpression of the BBX32 protein in soybean plant cells. Transgenicsoybean cells were regenerated into intact soybean plants and individualplants were selected from the population of independently transformedtransgenic plants that showed desirable molecular characteristics, suchas single copy of the transgene cassette at a single locus, theintegrity of the transgene cassette, absence of the construct backbonesequence, loss of the unlinked glyphosate resistance selection cassette.Furthermore, inverse PCR and DNA sequence analyses were performed todetermine the 5′ and 3′ insert-to-plant genome junctions, to confirm theorganization of the elements within the insert (FIG. 1), and todetermine the complete DNA sequence of the insert in soybean eventMON87712 (SEQ ID NO: 5). In addition, transgenic plants were screenedand selected for increased yield under field conditions. A soybean plantthat contains in its genome the BBX32 expression cassette of pMON83132is an aspect of the present invention.

Increased yield of a transgenic plant of the present invention can bemeasured in a number of ways, including test weight, seed number perplant, seed weight, seed number per unit area (i.e. seeds, or weight ofseeds, per acre), bushels per acre, tons per acre, or kilo per hectare.Expression of the BBX32 protein in plants comprising event MON87712leads to an increase in yield brought about by the increased capacityfor growth and reproductive development. Increased yield results fromthe partitioning of growth in one or more of the yield parametersincluding more seeds per plot, larger seeds, more seeds per plant,and/or more plants per plot. Recombinant DNA used in this inventionprovides increased yield through an increase in source compounds whencompared to non-transgenic soybean with the same genetic background.These biochemical and metabolic changes are linked todiurnally-regulated pathways that are impacted by the activity of BBX32and include carbon metabolism and nitrogen metabolism.

Methods for producing a plant with increased yield comprising transgenicevent MON87712 are provided. Transgenic plants used in these methods maybe homozygous or heterozygous for the transgene. Progeny plants producedby these methods may be varietal or hybrid plants; may be grown fromseeds produced by a plant and/or from seed comprising event MON87712produced by a plant fertilized with pollen from a plant comprising eventMON87712; and may be homozygous or heterozygous for the transgene.Progeny plants may be subsequently self-pollinated to generate a truebreeding line of plants, i.e., plants homozygous for the transgene, oralternatively may be outcrossed, e.g., bred with another unrelatedplant, to produce a varietal or a hybrid seed or plant. As used herein,the term “zygosity” refers to the similarity of DNA at a specificchromosomal location (locus) in a plant. In the present invention, theDNA specifically refers to the transgene insert along with the junctionsequence (event DNA). A plant is homozygous if the transgene insert withthe junction sequence is present at the same location on each chromosomeof a chromosome pair (2 alleles). A plant is considered heterozygous ifthe transgene insert with the junction sequence is present on only onechromosome of a chromosome pair (1 allele). A wild-type plant is nullfor the event DNA.

A plant with increased yield may be produced by sexually crossing aplant comprising event MON87712 comprising a polynucleotide having thenucleotide sequence of consecutive nucleotides of SEQ ID NO: 1, apolynucleotide having the nucleotide sequence of consecutive nucleotidesof SEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7,and/or at least 51 consecutive nucleotides of SEQ ID NO: 8 with anotherplant and thereby producing seed, which is then grown into progenyplants. These progeny plants may be analyzed using diagnostic methods toselect for progeny plants that contain event MON87712 DNA or for progenyplants with increased yield. The other plant used may or may not betransgenic. The progeny plant and/or seed produced may be varietal orhybrid seed.

A plant with increased yield may be produced by selfing a plantcomprising event MON87712 comprising a polynucleotide having thenucleotide sequence of consecutive nucleotides of SEQ ID NO: 1, apolynucleotide having the nucleotide sequence of consecutive nucleotidesof SEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7,and/or at least 51 consecutive nucleotides of SEQ ID NO: 8 and therebyproducing seed, which is then grown into progeny plants. These progenyplants may then be analyzed using diagnostic methods to select forprogeny plants that comprise event MON87712 DNA.

Progeny plants and seeds encompassed by these methods and produced byusing these methods are distinct from other plants, for example, becausethe progeny plants and seeds are recombinant and as such created byhuman intervention; contain at least one allele that consists of thetransgenic DNA of the present invention; and/or contain a detectableamount of a polynucleotide sequence selected from the group consistingof consecutive nucleotides of SEQ ID NO: 1, consecutive nucleotides ofSEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7, andat least 51 consecutive nucleotides of SEQ ID NO: 8. A seed may beselected from an individual progeny plant, and so long as the seedcomprises SEQ ID NO: 1, SEQ ID NO: 2, at least 51 consecutivenucleotides of SEQ ID NO: 7, and/or at least 51 consecutive nucleotidesof SEQ ID NO: 8, it will be within the scope of the present invention.

The plants, progeny, seeds, plant cells, plant parts (such as pollen,ovule, pod, flower, root or stem tissue, and leaves), and commodityproducts of the present invention may be evaluated for DNA composition,gene expression, and/or protein expression. Such evaluation may be doneby using various methods such as PCR, sequencing, northern blotting,southern analysis, western blotting, immuno-precipitation, and ELISA orby using the methods of detection and/or the detection kits providedherein.

Methods of detecting the presence of compositions specific to eventMON87712 in a sample are provided. One method consists of detecting thepresence of DNA specific to and derived from a cell, a tissue, a seed, aplant or plant parts comprising event MON87712. The method provides fora template DNA sample to be contacted with a primer pair that is capableof producing an amplicon from event MON87712 DNA upon being subjected toconditions appropriate for amplification, particularly an amplicon thatcomprises SEQ ID NO: 1, SEQ ID NO: 2, at least 51 consecutivenucleotides of SEQ ID NO: 7, and/or at least 51 consecutive nucleotidesof SEQ ID NO: 8, or the complements thereof. The amplicon is producedfrom a template DNA molecule derived from event MON87712, so long as thetemplate DNA molecule incorporates the specific and unique nucleotidesequences of SEQ ID NO: 1, SEQ ID NO: 2, at least 51 consecutivenucleotides of SEQ ID NO: 7, and/or at least 51 consecutive nucleotidesof SEQ ID NO: 8. The amplicon may be single or double stranded DNA orRNA, depending on the polymerase selected for use in the production ofthe amplicon. The method provides for detecting the amplicon moleculeproduced in any such amplification reaction, and confirming within thesequence of the amplicon the presence of the nucleotides correspondingto SEQ ID NO: 1, SEQ ID NO: 2, at least 51 consecutive nucleotides ofSEQ ID NO: 7, and/or at least 51 consecutive nucleotides of SEQ ID NO:8, or the complements thereof. The detection of the nucleotidescorresponding to SEQ ID NO: 1, SEQ ID NO: 2, at least 51 consecutivenucleotides of SEQ ID NO: 7, and/or at least 51 consecutive nucleotidesof SEQ ID NO: 8, or the complements thereof in the amplicon aredeterminative and/or diagnostic for the presence of event MON87712specific DNA and thus biological material comprising event MON87712 inthe sample.

Another method is provided for detecting the presence of a DNA moleculecorresponding to SEQ ID NO: 3 or SEQ ID NO: 4 in a sample consisting ofmaterial derived from plant or plant tissue. The method consists of (i)obtaining a DNA sample from a plant, or from a group of differentplants, (ii) contacting the DNA sample with a DNA probe moleculecomprising the nucleotides as set forth in either SEQ ID NO: 1 or SEQ IDNO: 2, (iii) allowing the probe and the DNA sample to hybridize understringent hybridization conditions, and then (iv) detecting ahybridization event between the probe and the target DNA sample.Detection of the hybrid composition is diagnostic for the presence ofSEQ ID NO: 3 or SEQ ID NO: 4, as the case may be, in the DNA sample.Absence of hybridization is alternatively diagnostic of the absence ofthe transgenic event in the sample if the appropriate positive controlsare run concurrently. Alternatively, determining that a particular plantcontains either or both of the sequences corresponding to SEQ ID NO: 1or SEQ ID NO: 2, or the complements thereof, is determinative that theplant contains at least one allele corresponding to event MON87712.

It is thus possible to detect the presence of a nucleic acid molecule ofthe present invention by any well known nucleic acid amplification anddetection methods such as polymerase chain reaction (PCR), recombinasepolymerase amplification (RPA), or DNA hybridization using nucleic acidprobes. An event-specific PCR assay is discussed, for example, byTaverniers et al. (J. Agric. Food Chem., 53: 3041-3052, 2005) in whichan event-specific tracing system for transgenic maize lines Bt11, Bt176,and GA21 and for transgenic event RT73 is demonstrated. In this study,event-specific primers and probes were designed based upon the sequencesof the genome/transgene junctions for each event. Transgenic plant eventspecific DNA detection methods have also been described in U.S. Pat.Nos. 6,893,826; 6,825,400; 6,740,488; 6,733,974; 6,689,880; 6,900,014and 6,818,807.

DNA detection kits are provided. One type of kit contains at least oneDNA molecule of sufficient length of contiguous nucleotides of SEQ IDNO: 3, SEQ ID NO: 5, or SEQ ID NO: 6 to function as a DNA primer orprobe specific for detecting the presence of DNA derived from transgenicevent MON87712 in a sample. The DNA molecule being detected with the kitcomprises contiguous nucleotides of the sequence as set forth in SEQ IDNO: 1 or at least 51 consecutive nucleotides of SEQ ID NO: 7.Alternatively, the kit may contain at least one DNA molecule ofsufficient length of contiguous nucleotides of SEQ ID NO: 4, SEQ ID NO:5, or SEQ ID NO: 6 to function as a DNA primer or probe specific fordetecting the presence of DNA derived from transgenic event MON87712 ina sample. The DNA molecule being detected with the kit comprisescontiguous nucleotides as set forth in SEQ ID NO: 2 or at least 51consecutive nucleotides of SEQ ID NO: 8.

An alternative kit employs a method in which the target DNA sample iscontacted with a primer pair as described above, then performing anucleic acid amplification reaction sufficient to produce an ampliconcomprising the consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, atleast 51 consecutive nucleotides of SEQ ID NO: 7, or at least 51consecutive nucleotides of SEQ ID NO: 8. Detection of the amplicon anddetermining the presence of the consecutive nucleotides of SEQ ID NO: 1,SEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7, or atleast 51 consecutive nucleotides of SEQ ID NO: 8, or the complementsthereof within the sequence of the amplicon is diagnostic for thepresence of event MON87712 specific DNA in a DNA sample.

A DNA molecule sufficient for use as a DNA probe is provided that isuseful for determining, detecting, or for diagnosing the presence oreven the absence of DNA specific and unique to event MON87712 DNA in asample. The DNA molecule contains the consecutive nucleotides of SEQ IDNO: 1, or the complement thereof, the consecutive nucleotides of SEQ IDNO: 2, or the complement thereof, at least 51 consecutive nucleotides ofSEQ ID NO: 7, or the complement thereof, or at least 51 consecutivenucleotides of SEQ ID NO: 8, or the complement thereof.

Nucleic acid amplification can be accomplished by any of the variousnucleic acid amplification methods known in the art, including thermaland isothermal amplification methods. The sequence of the heterologousDNA insert, junction sequences, or flanking sequences from eventMON87712 (with representative seed samples comprising event MON87712deposited as ATCC PTA-10296) can be verified by amplifying suchsequences from the event using primers derived from the sequencesprovided herein followed by standard DNA sequencing of the amplicon orof the cloned DNA.

The amplicon produced by these methods may be detected by a plurality oftechniques. One such method is Genetic Bit Analysis (Nikiforov, et al.Nucleic Acid Res. 22:4167-4175, 1994) where a DNA oligonucleotide isdesigned which overlaps both the adjacent flanking genomic DNA sequenceand the inserted DNA sequence. The oligonucleotide is immobilized inwells of a microwell plate. Following thermal amplification of theregion of interest (using one primer in the inserted sequence and one inthe adjacent flanking genomic sequence), a single-stranded amplicon canbe hybridized to the immobilized oligonucleotide and serve as a templatefor a single base extension reaction using a DNA polymerase and labelledddNTPs specific for the expected next base. Readout may be fluorescentor ELISA-based. Detection of a fluorescent or other signal indicates thepresence of the insert/flanking sequence due to successfulamplification, hybridization, and single base extension.

Another method is the Pyrosequencing technique as described by Winge(Innov. Pharma. Tech. 00:18-24, 2000). In this method an oligonucleotideis designed that overlaps the adjacent genomic DNA and insert DNAjunction. The oligonucleotide is hybridized to a single-strandedamplicon from the region of interest (one primer in the insertedsequence and one in the flanking genomic sequence) and incubated in thepresence of a DNA polymerase, ATP, sulfurylase, luciferase, apyrase,adenosine 5′ phosphosulfate and luciferin. ddNTPs are added individuallyand the incorporation results in a light signal which is measured. Alight signal indicates the presence of the transgene insert/flankingsequence due to successful amplification, hybridization, and single ormulti-base extension.

Fluorescence Polarization as described by Chen, et al., (Genome Res.9:492-498, 1999) is a method that can be used to detect the amplicon.Using this method an oligonucleotide is designed which overlaps thegenomic flanking and inserted DNA junction. The oligonucleotide ishybridized to single-stranded amplicon from the region of interest (oneprimer in the inserted DNA and one in the flanking genomic DNA sequence)and incubated in the presence of a DNA polymerase and afluorescent-labeled ddNTP. Single base extension results inincorporation of the ddNTP. Incorporation can be measured as a change inpolarization using a fluorometer. A change in polarization indicates thepresence of the transgene insert/flanking sequence due to successfulamplification, hybridization, and single base extension.

TAQMAN® (PE Applied Biosystems, Foster City, Calif.) may also be used todetect and/or quantifying the presence of a DNA sequence using theinstructions provided by the manufacturer. Briefly, a FREToligonucleotide probe is designed which overlaps the genomic flankingand insert DNA junction. The FRET probe and amplification primers (oneprimer in the insert DNA sequence and one in the flanking genomicsequence) are cycled in the presence of a thermostable polymerase anddNTPs. Hybridization of the FRET probe results in cleavage and releaseof the fluorescent moiety away from the quenching moiety on the FRETprobe. A fluorescent signal indicates the presence of theflanking/transgene insert sequence due to successful amplification andhybridization.

Molecular Beacons have been described for use in sequence detection asdescribed in Tyangi, et al. (Nature Biotech. 14:303-308, 1996). Briefly,a FRET oligonucleotide probe is designed that overlaps the flankinggenomic and insert DNA junction. The unique structure of the FRET proberesults in it containing secondary structure that keeps the fluorescentand quenching moieties in close proximity. The FRET probe andamplification primers (one primer in the insert DNA sequence and one inthe flanking genomic sequence) are cycled in the presence of athermostable polymerase and dNTPs. Following successful amplification,hybridization of the FRET probe to the target sequence results in theremoval of the probe secondary structure and spatial separation of thefluorescent and quenching moieties, which leads to the production of afluorescent signal. The fluorescent signal indicates the presence of theflanking/transgene insert sequence due to successful amplification andhybridization.

Other described methods, such as, microfluidics (US Patent PublicationNo. 2006068398, U.S. Pat. No. 6,544,734) provide methods and devices toseparate and amplify DNA samples. Optical dyes are used to detect andmeasure specific DNA molecules (WO/05017181). Nanotube devices(WO/06024023) that comprise an electronic sensor for the detection ofDNA molecules or nanobeads that bind specific DNA molecules and can thenbe detected.

DNA detection kits can be developed using the compositions disclosedherein and the methods well known in the art of DNA detection. The kitsare useful for the identification of event MON87712 in a sample and canbe applied to methods for breeding plants containing the appropriateevent DNA. The kits may contain DNA primers or probes that are similaror complementary to SEQ ID NO: 1-6, or fragments or complements thereof.

The kits and detection methods of the present invention are thereforeuseful for, among other things, identifying event MON87712, selectingplant varieties or hybrids comprising event MON87712, detecting thepresence of DNA derived from event MON87712 in a sample, and monitoringsamples for the presence and/or absence of event MON87712 or plants,plant parts or commodity products comprising event MON87712.

The following examples are included to demonstrate examples of certainembodiments of the invention. It should be appreciated by those of skillin the art that the techniques disclosed in the examples that followrepresent approaches the inventors have found function well in thepractice of the invention, and thus can be considered to constituteexamples of preferred modes for its practice. However, those of skill inthe art should, in light of the present disclosure, appreciate that manychanges can be made in the specific embodiments that are disclosed andstill obtain a like or similar result without departing from the spiritand scope of the invention.

EXAMPLES Example 1: Transformation of Soybean with pMON83132 and EventSelection

This example describes transformation and generation of transgenicsoybean events, and selection of event MON87712.

An Agrobacterium-mediated transformation method was used to transformsoybean cells with a binary construct designated as pMON83132. Theconstruct contains two plant transformation cassettes or T-DNAs. Eachcassette is flanked by right border and left border sequences. Thetransgenic insert as set forth in SEQ ID NO: 5 comprises atransformation cassette containing an enhanced 35S promoter fromcauliflower mosaic virus (CaMV), operably linked to a DNA moleculeencoding BBX32 protein, operably linked to the 3′ untranslated regionfrom fiber protein E6 gene of Gossypium barbadense (cotton). The secondtransformation cassette contains a chimeric promoter consisting ofenhancer sequences from the promoter of the figwort mosaic virus (FMV)35S RNA combined with the promoter of the EF-1α gene from Arabidopsisthaliana that encodes elongation factor EF-1 alpha. It also contains aleader (exon 1) and intron with flanking splice sites of the ArabidopsisEF-1α gene, operably linked to a DNA molecule encoding a chloroplasttransit peptide (CTP2) from Arabidopsis EPSP synthase, fused to a codonmodified coding sequence of the aroA gene from the Agrobacterium sp.strain CP4 encoding the CP4 EPSPS protein, operably linked to a 3′untranslated region of the RbcS2 gene from Pisum sativum. The CP4 aroAgene confers tolerance to glyphosate, and was used as a selectablemarker. Table 1 is a summary of the genetic elements in pMON83132.

After transformation with construct pMON83132, transformed cells wereallowed to grow and multiply on media containing glyphosate. Plants wereregenerated from surviving cells. A total of 4944 independent R0transformation events were produced and characterized by detailedmolecular analyses. Events were screened by quantitative PCR and RT-PCR(TAQMAN®) for insert number (number of integration sites within thesoybean genome), copy number (the number of copies of the T-DNA withinone locus), the integrity of the inserted cassette and the absence ofbackbone sequence, and expression of the AtBBX32 transgene transcript.Events with undesirable molecular characteristics, such as presence ofmultiple copies of the transgene and/or molecular complexity of theinsert, the presence of the transformation vector backbone sequence wereeliminated. Furthermore, linkage Southern analysis was done to removeevents where the BBX32 expression cassette was linked to the CP4selectable marker cassette. A total of 284 events met the stringentmolecular selection criteria based on the analyses described above.Three additional events were further eliminated due to unavailability ofhomozygous seed stocks, resulting in 281 events being advanced to R1generation.

TABLE 1 Summary of the genetic elements in pMON83132. Location inGenetic Construct Element pMON83132 Function (Reference) T-DNA I B¹-LeftBorder  1-442 DNA region from Agrobacterium tumefaciens containing theleft border sequence used for transfer of the T-DNA (Barker et al.,Plant Mol. Biol. 2: 335- 350, 1983). P²-e35S  511-1123 Promoter for thecauliflower mosaic virus (CaMV) 35S RNA (Odell et al., Nature 313:810-812, 1985) containing the duplicated enhancer region (Kay et al.,Science 236: 1299-1302, 1987) that directs transcription in plant cells.CS³-BBX32 1148-1825 Coding sequence of the BBX32 gene from Arabidopsisthaliana encoding a zinc finger protein (B-box type) (Khanna et al.,Plant Cell 21: 3416- 3420, 2009) which modulates aspects of diurnalbiology (Holtan et al., submitted). T⁴-E6 1840-2154 3′ UTR region of theE6 gene of Gossypium barbadense (cotton) that encodes a fiber proteininvolved in early fiber development (John, Plant Mol Biol. 30: 297-306,1996) which functions to direct polyadenylation of mRNA. B-Right Border2193-2549 DNA region from Agrobacterium tumefaciens containing the rightborder sequence used for transfer of the T-DNA (Depicker et al., J. ofMol. and Applied Genetics 1: 561-573, 1982; Zambryski et al., J. of Mol.and Applied Genetics 1: 361-370, 1982). T-DNA II B-Right Border2721-3077 DNA region from Agrobacterium tumefaciens containing the rightborder sequence used for transfer of the T-DNA (Depicker et al., J. ofMol. and Applied Genetics 1: 561-573, 1982; Zambryski et al., J. of Mol.and Applied Genetics 1: 361-370, 1982). P-FMV/EF-1α 3100-4139 Chimericpromoter consisting of enhancer sequences from the promoter of theFigwort Mosaic virus (FMV) 35S RNA (Richins et al., Nucleic AcidsResearch 15: 8451-8466, 1987) combined with the promoter from theEF-1αgene of Arabidopsis thaliana that encodes elongation factor EF-1alpha (Axelos et al., Molecular and General Genetics 219: 106-112,1989). It is associated with constitutive expression of the gene.L⁵-EF-1α 4140-4185 Leader (exon 1) of the EF-1αgene from Arabidopsisthaliana that encodes elongation factor EF-1 alpha (Axelos et al.,Molecular and General Genetics 219: 106-112, 1989), which enhances geneexpression. I⁶-EF-1α 4186-4807 Intron with flanking splice sites of theEF-1α gene from Arabidopsis thaliana that encodes elongation factor EF-1alpha (Axelos et al., Molecular and General Genetics 219: 106-112,1989), that enhances gene expression. TS⁷-CTP2 4817-5044 Targetingsequence of the ShkG gene from Arabidopsis thaliana encoding EPSPScontaining the transit peptide region) that directs transport of theEPSPS protein to the chloroplast (Klee et al., Molecular and GeneralGenetics 210: 437-442, 1987). CS-modified cp4 5045-6412 Codon modifiedcoding sequence of the aroA gene epsps from the Agrobacterium sp. strainCP4 encoding the CP4 EPSPS protein (Padgette et al., 1996, inHerbicide-Resistant Crops: Agricultural, Economic, Environmental,Regulatory, and Technological Aspects, S. O. Duke, (ed.), Pages 53-84,CRC Press, Boca Raton, FL). T-E9 6419-7061 3′ nontranslated region ofthe RbcS2 gene from Pisum sativum (pea) encoding the Rubisco smallsubunit, which functions to direct polyadenylation of the mRNA (Coruzziet al., EMBO J. 3: 1671-1679, 1984). B-Left Border 7076-7486 DNA regionfrom Agrobacterium tumefaciens containing the left border sequence usedfor transfer of the T-DNA (Barker et al., Plant Mol. Biol. 2: 335- 350,1983). Vector Backbone aadA 7582-8470 Bacterial promoter, codingsequence, and 3′ UTR for an aminoglycoside-modifying enzyme, 3″(9)-O-nucleotidyltransferase from the transposon Tn7 (Fling et al., NucleicAcids Research 13: 7095-7106, 1985) that confers spectinomycin andstreptomycin resistance. OR⁸-ori-pBR322 9001-9589 Origin of replicationfrom pBR322 for maintenance of plasmid in E. coli (Sutcliffe, 1979,Complete nucleotide sequence of the Escherichia coli plasmid pBR322.Pages 77-90 in Cold Spring Harbor Symposia on Quantitative Biology, ColdSpring Harbor, NY, Cold Spring Harbor Laboratory Press). CS-rop10017-10208 Coding sequence for repressor of primer protein derived fromthe ColE1 plasmid for maintenance of plasmid copy number in E. coli(Giza and Huang, Gene 78: 73-84, 1989). OR-ori V 10946-11342 Origin ofreplication from the broad host range plasmid RK2 for maintenance ofplasmid in Agrobacterium (Stalker et al., Molecular and General Genetics181: 8-12, 1981). ¹B, Border ²P, Promoter ³CS, Coding Sequence ⁴T,Transcription Termination Sequence ⁵L, Leader ⁶I, Intron ⁷TS, TargetingSequence ⁸OR, Origin of Replication

At the R1 generation, events underwent segregation analysis to selectonly homozygous and heterozygous plants going forward. In addition, CP4linkage analysis, TAQMAN® analysis and BBX32 expression analysis wereperformed to remove events where CP4 was linked to the BBX32 transgene,or where the sequence of the OriV origin of replication (backbone) wasdetected, or where BBX32 transgene expression level was undesirable,resulting in 121 events remaining. Southern hybridization analysis usingdifferent probes further eliminated 42 events that were not single copy,or single insert or not backbone-free. Probes included the intact BBX32coding region and its respective promoter, the polyadenylation sequenceand the plasmid backbone. An additional 2 events were stopped due tolack of sufficient seeds to advance, leaving 77 events to carry forwardto the R2 generation.

At the R2 generation, an additional 9 events were dropped due to 1) lackof sufficient seed for the R2 nursery (1 event), or 2) undesirableagronomic phenotypes (2 events), or 3) BBX32 expression level or theabsence of the representative homozygous lines as determined by Invader®(Third Wave Technologies, Inc., Madison, Wis.) (6 events). As a result,68 events were carried forward for year 1 yield field test in the US.Based upon yield performance, 16 events were advanced to the secondseason field testing in South America, from which, 9 events wereselected for further yield testing. Meta-analysis of yield data from 4seasons of field trials in the United States or South America, each withmultiple locations (Table 2), showed that one progeny line designated ascomprising event MON87712 was the highest yielding event (FIG. 2).

TABLE 2 Details of the four season field testing for yield. YearGeography # Locations 2007 United States 18 2007-2008 South America 142008 United States 24 2008-2009 South America 14

Example 2: Isolation of Flanking Sequences Using Inverse PCR AndIdentification of Flanking Sequences by Sequencing

This example describes isolation of the soybean genomic DNA sequencesflanking the transgenic DNA insert using inverse PCR for event MON87712,and identification of the flanking genomic sequences by sequencing.

Sequences flanking the T-DNA insertion in event MON87712 were determinedusing inverse PCR as described in Ochman et al., 1990 (PCR Protocols: Aguide to Methods and Applications, Academic Press, Inc.). Plant genomicDNA was isolated from both wild-type A3525 and the transgenic line fromtissue grown under greenhouse conditions. Frozen leaf tissue was groundwith a mortar and a pestle in liquid nitrogen or by mechanical grinding,followed by DNA extraction using methods known in the art. This methodcan be modified by one skilled in the art to extract DNA from anytissue, including, but not limited to seed.

An aliquot of DNA from each sample was digested with restrictionendonucleases selected based on restriction analysis of the transgenicDNA. After self-ligation of the restriction fragments, PCR amplificationwas performed using primers designed from the transgenic sequence thatwould amplify sequences extending away from the 5′ and 3′ ends of thetransgenic DNA. A variety of Taq polymerases and amplification systemsmay be used. Table 3 shows an example of PCR amplification for flankingsequence isolation using Phusion High Fidelity DNA Polymerase (Cat. No.F531S or F531L, New England Biolabs), and Thermalcyclers AppliedBiosystems GeneAmp 9700, ABI 9800 Fast Thermal Cycler and MJ Opticon.

TABLE 3 An example of inverse PCR amplification for flanking sequenceisolation. Volume Component PCR master mix (per 2.9 μl Water reaction)0.05 μl Primer 1 (100 μM) 0.05 μl Primer 1 (100 μM) 5.0 μl 2X PhusionTaq 2.0 μl ligated DNA 10 μl Total Step Condition DNA amplification in afast 1 98° C. 30 sec thermocycler 2 98° C. 5 sec 3 60° C. 10 sec 4 72°C. 2 min 5 Go to step 2 30 times 6 72° C. 4 min 7 10° C. forever 8 End

PCR products were separated by agarose gel electrophoresis and purifiedusing a QIAGEN gel purification kit (Qiagen, Valencia, Calif.). Thesubsequent products were sequenced directly using standard sequencingprotocols. Using these two methods, the 5′ flanking sequence, whichextends into the left border sequence of the integrated DNA insertincluding the BBX32 expression cassette, was identified and is presentedas SEQ ID NO: 3 ([C] of FIG. 1). The 3′ flanking sequence, which extendsinto the right border sequence of the integrated DNA insert includingthe BBX32 expression cassette, was identified and is presented as SEQ IDNO: 4 ([D] of FIG. 1). The transgenic DNA integrated into the soybeangenomic DNA is presented as SEQ ID NO: 5 ([E] of FIG. 1).

The isolated sequences were compared to the T-DNA sequence to identifythe flanking sequences and the co-isolated T-DNA fragments. Confirmationof the presence of the expression cassette was achieved by PCR withprimers designed based upon the deduced flanking sequence data and theknown T-DNA sequence. The wild type sequence corresponding to the sameregion in which the T-DNA was integrated in the transformed line wasisolated using primers designed from the flanking sequences in MON87712.The flanking sequences in MON87712 and the wild type sequence wereanalyzed against multiple nucleotide and protein databases. Thisinformation was used to examine the relationship of the transgene to theplant genome and to look at the insertion site integrity. The flankingsequence and wild type sequences were used to design primers for TAQMAN®endpoint assays used to identify the events and determine zygosity asdescribed in Example 3.

Example 3: Event-Specific Endpoint TAQMAN® And Zygosity Assays

This example describes an event-specific endpoint TAQMAN® thermalamplification method for identification of event MON87712 DNA in asample.

Examples of conditions useful with the event MON87712-specific endpointTAQMAN® method are described in Table 4 and Table 5. The DNA primersused in the endpoint assay are primers SQ3983 (SEQ ID NO: 16) andSQ22982 (SEQ ID NO: 17) and the 6-FAM™ labeled oligonucleotide probe isPB10453 (SEQ ID NO: 18). 6FAM™ is a fluorescent dye product of AppliedBiosystems (Foster City, Calif.) attached to the DNA probe. For TAQMAN®MGB (Minor Groove Binding) probes, the 5′ exonuclease activity of TaqDNA polymerase cleaves the probe from the 5′-end, between thefluorophore and quencher. When hybridized to the target DNA strand,quencher and fluorophore are separated enough to produce a fluorescentsignal.

Primers SQ3983 (SEQ ID NO: 16) and SQ22982 (SEQ ID NO: 17), when used asdescribed with probe PB10453 (SEQ ID NO: 18), produce an amplicon thatis diagnostic for event MON87712 DNA. The analysis includes a positivecontrol from soybean known to contain event MON87712 DNA, a negativecontrol from non-transgenic soybean and a negative control that containsno template DNA.

These assays are optimized for use with Applied Biosystems GeneAmp PCRSystem 9700, ABI 9800 Fast Thermal Cycler and MJ Opticon. Other methodsand apparatus known to those skilled in the art may be used to produceamplicons that identify the event MON87712 DNA.

TABLE 4 Soybean MON87712 Event-Specific Endpoint TAQMAN ® PCR ConditionsStep Reagent Volume Comments 1 18 megohm water adjust for final volumeof 10 μl 2 2X Universal Master Mix 5.0 μl 1X final concentration (dNTPs,enzyme and buffer) of dNTPs, enzyme and buffer 3 Event Primers SQ3983and 0.5 μl 1.0 μM final SQ22982 Mix (resuspended concentration in 18megohm water to a concentration of 20 μM for each primer) Example: In amicrocentrifuge tube, the following are added to achieve 500 μl at afinal concentration of 20 μM: 100 μl of Primer SQ22982 at aconcentration of 100 μM 100 μl of Primer SQ3983 at a concentration of100 μM 300 μl of 18 megohm water 4 Event 6-FAM ™ MGB Probe 0.2 μl 0.2 μMfinal PB10453 concentration (resuspended in 18 megohm water to aconcentration of 10 μM) 5 Internal control Primer-1 and 0.5 μl 1 μMfinal internal control Primer-2 Mix concentration (resuspended in 18megohm water to a concentration of 20 μM for each primer) 6 Internalcontrol VIC ™ probe 0.2 μl 0.2 μM final (resuspended in 18 megohmconcentration water to a concentration of 10 μM) 7 Extracted DNA(template): 3.0 μl 1. Leaf or seed samples to be analyzed 2. Negativecontrol (non-transgenic DNA) 3. Negative water control (no templatecontrol) 4. Positive control (MON87712DNA)

TABLE 5 Endpoint TAQMAN ® thermocycler conditions Cycle No. Settings 150° C. 2 minutes 1 95° C. 10 minutes 10 95° C. 15 seconds 64° C. 1minute (−1° C./cycle) 30 95° C. 15 seconds 54° C. 1 minute 1 10° C.Forever

The following example describes an event-specific endpoint TAQMAN®thermal amplification method developed to determine the zygosity ofevent MON87712 in a sample. A zygosity assay is useful for determiningif a plant comprising an event is homozygous for the event DNA, that iscomprising the exogenous DNA at the same location on each chromosome ofa chromosomal pair; or heterozygous for an event DNA, that is comprisingthe exogenous DNA on only one chromosome of a chromosomal pair; or isnull for the event DNA, that is wild type. A set of three primers (SEQID NO: 10, 13 and 11 or 15), a 6FAM™ labeled probe (SEQ ID NO: 12) and aVIC™ labeled probe (SEQ ID NO: 14) were used in the assays (Table 8).6FAM™ and VIC™ are fluorescent dye products of Applied Biosystems(Foster City, Calif.) attached to the DNA probes. Primer SEQ ID NO: 11or 15 hybridizes and extends specifically from the inserted exogenousDNA; primer SEQ ID NO: 10 hybridizes and extends specifically from theDNA flanking the 3′ side of the inserted exogenous DNA; primer SEQ IDNO: 13 hybridizes and extends specifically from the wild-type DNAcorresponding to a region that was deleted during T-DNA insertion in thetransgenic event MON87712. These primers are diagnostic. In thisexample, primers SEQ ID NO: 10 and SEQ ID NO: 11 or 15 and the6FAM™-labeled oligonucleotide probe SEQ ID NO: 12 produce a DNA ampliconrevealed by the liberation of a fluorescent signal from probe SEQ ID NO:12, which is diagnostic for event MON87712 DNA, indicating a copy of theinserted transgenic DNA present in the genomic DNA. Primers SEQ ID NO:10 and SEQ ID NO: 13 and the VIC™-labeled oligonucleotide probe SEQ IDNO: 14 produce an amplicon revealed by the liberation of a fluorescentsignal from probe SEQ ID NO: 14, which is diagnostic for the wild typeallele, indicating no copy of the inserted exogenous DNA present in thegenomic DNA. When the three primers and two probes are mixed together ina PCR reaction with DNA extracted from a plant, release of a fluorescentsignal only from the 6FAM™-labeled oligonucleotide probe (SEQ ID NO: 12)is indicative of and diagnostic of a plant homozygous for eventMON87712. When the three primers and two probes are mixed together in aPCR reaction with DNA extracted from a plant, release of a fluorescentsignal from both the 6FAM™-labeled oligonucleotide probe SEQ ID NO: 12and the VIC™-labeled oligonucleotide probe SEQ ID NO: 14 is indicativeof and diagnostic of a plant heterozygous for event MON87712. When thethree primers and two probes are mixed together in a PCR reaction withDNA extracted from a plant, release of a fluorescent signal from onlythe VIC™-labeled oligonucleotide probe SEQ ID NO: 14 is indicative ofand diagnostic of a plant null for event MON87712, i.e. wild type. Theassays also include a positive control from soybean containing eventMON87712 DNA, a negative control from non-transgenic soybean and anegative control that contains no template DNA.

Table 6 and Table 7 provide examples of conditions used for zygosityassay for event MON87712 in a sample. These assays are optimized for usewith an Applied Biosystems GeneAmp PCR System 9700. The allelicdetection was optimized with a 7900HT Sequence Detection System byApplied Biosystems, or a PHERAstar by BMG Labtech. Other methods andapparatus known to those skilled in the art that produce amplicons thatidentify the event MON87712 DNA is within the skill of the art.

TABLE 6 Soybean MON87712 Event-Specific Zygosity Endpoint TAQMAN ® PCRStep Details 1 Prepare the following: 200 μM Primers 100 μM Event6-FAM ™ MGB* probe 100 μM Wild type VIC ™ MGB probe 50:50 2X ABIuniversal master mix:2X AbGene universal master mix 0.1X TE 2 Preparethe following 80X Primer + probe stock: 100 μM Event 6-FAM ™ MGB Probe16 μl 100 μM Wild type VIC ™ MGB probe 16 μl 200 μM Wild type forwardprimer 8.5 μl 200 μM Event forward primer 8.5 μl 200 μM reverse primer17 μl 3 Dilute the 80X Primer + Probe stock to 10X working dilution with0.1X TE 4 Prepare the following reaction solution (per reaction): 50:502X ABI universal master mix:2X 3 μl AbGene universal master mix 10XPrimer + Probe working dilution 0.6 μl Extracted DNA (template): 2.4μl 1) Samples to be analyzed 2) Negative control (non-transgenic DNA) 3)Negative water control (no template control) 4) Positive controlhomozygous MON87712 DNA 5) Positive control heterozygous MON87712 DNA*MGB: Minor Groove Binding

TABLE 7 Zygosity Endpoint TAQMAN ® Thermocycler Conditions Cycle No.Settings 1 95° C. 10 minutes 40 92° C. 15 seconds 64° C. 1 Minute 1 10°C. Forever

TABLE 8  Examples of Primer And Probe Combinations Usedfor Zygosity Assays Combi- SEQ nation Type Direction ID NO Sequences 1Primers Reverse 10 GTTTTACAATTACCTCG TTTAAGTAAATCA Forward 13CTATTATTTGCTATAAG TATTTGATGTAAGAA Probe Wild type 14 VIC- alleleTTGTATTAATAACAAA AAATTG Primers Reverse 10 GTTTTACAATTACCTCGTTTAAGTAAATCA Forward 11 CATTCTCGAGCAGGAC CTGCAGAA Probe MON87712 126FAM- allele AACACTGATAGTTTAA ACTGAAG 2 Primers Reverse 10GTTTTACAATTACCTCG TTTAAGTAAATCA Forward 13 CTATTATTTGCTATAAGTATTTGATGTAAGAA Probe Wild type 14 VIC- allele TTGTATTAATAACAAA AAATTGPrimers Reverse 10 GTTTTACAATTACCTCG TTTAAGTAAATCA Forward 15GTACATTCTCGAGCAG GACCTGCA Probe MON87712 12 6FAM- alleleAACACTGATAGTTTAA ACTGAAG

Example 4: Identification of Event MON87712 in Any MON87712 ContainingBreeding Event

This example describes how one may identify the MON87712 event withinprogeny of any breeding event containing MON87712 soybean.

Event DNA primer pairs are used to produce an amplicon diagnostic forsoybean event MON87712. An amplicon diagnostic for MON87712 comprises atleast one junction sequence, provided herein as SEQ ID NO: 1, SEQ ID NO:2, at least 51 consecutive nucleotides of SEQ ID NO: 7, or at least 51consecutive nucleotides of SEQ ID NO: 8, or the complements thereof ([A]or [B], respectively as illustrated in FIG. 1). SEQ ID NO: 1 ([A] ofFIG. 1) is a nucleotide sequence corresponding to the junction of the 5′flanking sequence (positions 3495 through 3516 of SEQ ID NO: 3 [C], seeFIG. 1) and the left border of the integrated DNA insert (positions 1through 11 of SEQ ID NO: 5 [E], see FIG. 1). SEQ ID NO: 2 ([B] ofFIG. 1) is a nucleotide sequence corresponding to the junction of theright border of the integrated DNA insert (positions 2004 through 2014of SEQ ID NO: 5 [E], see FIG. 1) and the 3′ flanking sequence (positions90 through 111 of SEQ ID NO: 4 [D], see FIG. 1). SEQ ID NO: 7 ([A] ofFIG. 1) is a nucleotide sequence corresponding to the junction of the 5′flanking sequence (positions 3456 through 3555 of SEQ ID NO: 3 [C], seeFIG. 1) and the left border of the integrated DNA insert (positions 1through 50 of SEQ ID NO: 5 [E], see FIG. 1). SEQ ID NO: 8 ([B] ofFIG. 1) is a nucleotide sequence corresponding to the junction of theright border of the integrated DNA insert (positions 1965 through 2014of SEQ ID NO: 5 [E], see FIG. 1) and the 3′ flanking sequence (positions51 through 150 of SEQ ID NO: 4 [D], see FIG. 1).

Event primer pairs that produce an amplicon diagnostic for MON87712include primer pairs designed using the flanking sequences (SEQ ID NO: 3and SEQ ID NO: 4) and the integrated transgenic DNA sequence (SEQ ID NO:5). To acquire a diagnostic amplicon in which at least 15 nucleotides ofSEQ ID NO: 1 is found, one would design a forward primer based upon SEQID NO: 3 from bases 1 through 3494 and a reverse primer based upon theinserted expression cassette DNA sequence, SEQ ID NO: 5 from positions12 through 2014, in which the primers are of sufficient length ofcontiguous nucleotides to specifically hybridize to SEQ ID NO: 3 and SEQID NO: 5. To acquire a diagnostic amplicon in which at least 15nucleotides of SEQ ID NO: 2 is found, one would design a forward primerbased upon the inserted expression cassette DNA sequence, SEQ ID NO: 5,from positions 1 through 2003 and a reverse primer based upon the 3′flanking sequence, SEQ ID NO: 4, from bases 12 through 2065, in whichthe primers are of sufficient length of contiguous nucleotides tospecifically hybridize to SEQ ID NO: 4 and SEQ ID NO: 5. For practicalpurposes, one should design primers that produce amplicons of a limitedsize range, for example, between 100 to 1000 bases. Smaller sized(shorter nucleotide length) amplicons in general may be more reliablyproduced in PCR reactions, allow for shorter cycle times and be easilyseparated and visualized on agarose gels or adapted for use in endpointTAQMAN®-like assays. In addition, amplicons produced using primer pairscan be cloned into vectors, propagated, isolated and sequenced or can besequenced directly with methods well established in the art. Any primerpair derived from the combination of SEQ ID NO: 3 and SEQ ID NO: 5 orthe combination of SEQ ID NO: 4 and SEQ ID NO: 5 that are useful in aDNA amplification method to produce an amplicon diagnostic for MON87712,plants comprising MON87712 or progeny thereof is an aspect of thepresent invention. Any single isolated DNA primer molecule comprising atleast 11 contiguous nucleotides of SEQ ID NO: 3, or its complement thatis useful in a DNA amplification method to produce an amplicondiagnostic for MON87712, plants comprising MON87712 or progeny thereofis an aspect of the present invention. Any single isolated DNA primermolecule comprising at least 11 contiguous nucleotides of SEQ ID NO: 4,or its complement that is useful in a DNA amplification method toproduce an amplicon diagnostic for MON87712, plants comprising MON87712or progeny thereof is an aspect of the present invention. Any singleisolated DNA primer molecule comprising at least 11 contiguousnucleotides of SEQ ID NO: 5, or its complement that is useful in a DNAamplification method to produce an amplicon diagnostic for MON87712,plants comprising MON87712 or progeny thereof is an aspect of thepresent invention.

An example of the amplification conditions for this analysis isillustrated in Table 4 and Table 5 (Example 3). However, anymodification of these methods or the use of DNA primers homologous orcomplementary to SEQ ID NO: 3, SEQ ID NO: 4 or DNA sequences of thetransgenic insert (SEQ ID NO: 5) of event MON87712 that produce anamplicon diagnostic for MON87712 is within the scope of the presentdisclosure. A diagnostic amplicon comprises a DNA molecule homologous orcomplementary to at least one transgene/genomic junction DNA (SEQ ID NO:1, SEQ ID NO: 2, at least 51 consecutive nucleotides of SEQ ID NO: 7, orat least 51 consecutive nucleotides of SEQ ID NO: 8, or the complementsthereof), or a substantial portion thereof.

An analysis for event MON87712 DNA in a sample should include a positivecontrol from event MON87712, a negative control from a soybean plantthat does not contain event MON87712, for example, but not limited towild type control, and a negative control that contains no soybeangenomic DNA. A primer pair that will amplify an endogenous soybean DNAmolecule will serve as an internal control for the DNA amplificationconditions. Additional primer sequences can be selected from SEQ ID NO:3, SEQ ID NO: 4, or SEQ ID NO: 5 by those skilled in the art of DNAamplification methods, and conditions selected for the production of anamplicon by the methods shown in Table 4 and Table 5 may differ, butresult in an amplicon diagnostic for event MON87712 DNA. The use ofthese DNA primer sequences with modifications to the methods of Table 4and Table 5 are within the scope of the invention. The amplicon producedby at least one DNA primer sequence derived from SEQ ID NO: 3, SEQ IDNO: 4, or SEQ ID NO: 5 that is diagnostic for MON87712 is an aspect ofthe invention.

DNA detection kits that contain at least one DNA primer derived from SEQID NO: 3, SEQ ID NO: 4, or SEQ ID NO: 5, that when used in a DNAamplification method, produces a diagnostic amplicon for MON87712,plants comprising MON87712 or progeny thereof is an aspect of theinvention. A soybean plant or seed, wherein its genomic DNA produces anamplicon diagnostic for MON87712 when tested in a DNA amplificationmethod is an aspect of the invention. The assay for the MON87712amplicon can be performed by using an Applied Biosystems GeneAmp PCRSystem 9700, ABI 9800 Fast Thermal Cycler and MJ Opticon, StratageneRobocycler, MJ Engine, Perkin-Elmer 9700 or Eppendorf MastercyclerGradient thermocycler or any other amplification system that can be usedto produce an amplicon diagnostic of MON87712.

A deposit of the soybean event MON87712 comprising seed disclosed aboveand recited in the claims, has been made under the Budapest Treaty withthe American Type Culture Collection (ATCC), 10801 University Boulevard,Manassas, Va. 20110. The date of deposit is Aug. 20, 2009 and the ATCCaccession number is PTA-10296. Upon issuance of a patent, allrestrictions upon the deposit will be removed, and the deposit isintended to meet all of the requirements of 37 C.F.R. §§ 1.801-1.809.The deposit will be maintained in the depository for a period of 30years, or 5 years after the last request, or for the effective life ofthe patent, whichever is longer, and will be replaced as necessaryduring that period.

Having illustrated and described the principles of the presentinvention, it should be apparent to persons skilled in the art that theinvention can be modified in arrangement and detail without departingfrom such principles. We claim all modifications that are within thespirit and scope of the appended claims.

1-15. (canceled)
 16. A hybrid soybean plant or seed wherein at least oneparent is derived from a soybean comprising event MON87712. 17-23.(canceled)
 24. A method of determining the zygosity of a soybean plantgenome comprising soybean event MON87712 DNA in a sample comprising: a)contacting the sample with three different primers that i) when usedtogether in a nucleic acid amplification reaction with soybean eventMON87712 DNA, produces a first amplicon that is diagnostic for soybeanevent MON87712; and ii) when used together in a nucleic acidamplification reaction with soybean genomic DNA other than eventMON87712 DNA, produces a second amplicon that is diagnostic for soybeanwild type genomic DNA other than event MON87712 DNA; b) performing anucleic acid amplification reaction; and c) detecting said firstamplicon and said second amplicon; wherein the presence of said firstand second amplicons is diagnostic of a genome heterozygous for eventMON87712 in said sample, and wherein the presence of only said firstamplicon is diagnostic of a genome homozygous for event MON87712 in saidsample.
 25. The method of claim 24, wherein the three different primerscomprise SEQ ID NO: 10, SEQ ID NO: 11 or 15, and SEQ ID NO:
 13. 26.(canceled)